- 13th International Workshop on Langerhans Cells

Transcription

LC 2013
13th International Workshop
on Langerhans Cells
October 10 -13, 2013
Amsterdam, The Netherlands
SCIENTIFIC PROGRAM
& ABSTRACTS
WELCOME
Ladies and gentlemen, friends and colleagues, Langerhans researchers all over the world, The fact that the Nobel Prize in Medicine 2011 was awarded to Ralph M. Steinman "for his discovery of the dendritic cell and its role in adaptive immunity" underlines the importance of dendritic cells. Dendritic cells, including epidermal skin Langerhans cells are considered to be master regulators for protective immunity and tolerance. Langerhans cell research is an established niche within research on dendritic cells, encompassing both fundamental and clinical immunology. The development of exiting new experimental models boosted interest in skin dendritic cells, also including dermal dendritic cells. All basic research on Langerhans cells and dermal dendritic cells over the last decades is beginning to pay off, as clinical opportunities become clear, such as intradermal vaccination. The tradition of having biennial International Langerhans Cell Workshops started in 1989 in Oslo, Norway, and we are happy to host the 13th workshop in Amsterdam. The rich cultural background of the capital city of The Netherlands, together with its many social opportunities, guarantees a successful meeting and the initiation of new collaborations and interactions between advanced and young researchers but also between researchers and industry. LC2013 will focus on the function of Langerhans cells and other dendritic cell subsets in skin and mucosa. The recent developments in the field will be highlighted with excellent invited speakers, state‐of‐the art oral presentations and poster sessions. We cordially invite researchers, dermatologists, and industry to actively participate and interact in the 13th International Workshop on Langerhans Cells in Amsterdam, The Netherlands. LC2013 provides the great opportunity to listen to international experts to hear their latest news and their opinions. The meeting will take place from Thursday, October 10th to Sunday, October 13th, 2013 at the Royal Tropical Institute, a beautiful monumental building and centrally located in the city of Amsterdam. The local organizers have been working hard to ensure that you will have a great time and plenty of new information to share with your colleagues when you return home. We thank all sponsors for their support without which this Workshop would not be possible. Welcome to Amsterdam ! Teunis Geijtenbeek, Esther de Jong & Marcel Teunissen Under the auspices of: Academic Medical Center University of Amsterdam Amsterdam The Netherlands 1 COMMITTEES
Local Organizing Committee
Marcel B.M. Teunissen, Amsterdam, The Netherlands
Esther C. de Jong, Amsterdam, The Netherlands
Teunis B.H. Geijtenbeek, Amsterdam, The Netherlands
International Organizing Committee
Björn Clausen, Mainz, Germany
Adelheid Elbe-Bürger, Vienna, Austria
Florent Ginhoux, Singapore
Sandrine Henri, Marseille, France
Juliana Idoyaga, New York, USA
Daniel Kaplan, Minneapolis, MN, USA
Eynav Klechevsky, St. Louis, MO, USA
Miriam Merad, New York, USA
Keisuke Nagao, Tokyo, Japan
Niki Romani, Innsbruck, Austria
Georg Stingl, Vienna, Austria
Patrizia Stoitzner, Innsbruck, Austria
Marcel Teunissen, Amsterdam, Netherlands
Mark Udey, Bethesda, MD, USA
James W. Young, New York
Past Organizers* - Past Advisory Committee
Paul Bergstresser*, Dallas, USA
Lasse R. Braathen*, Berne, Switzerland
Colette Dezutter-Dambuyant*, Lyon, France
Adelheid Elbe-Bürger*, Vienna, Austria
Giampiero Girolomoni*, Verona, Italy
Richard D. Granstein*, New York, USA
Mieke Mommaas*, Leiden, The Netherlands
Gwendalyn J. Randolph, St. Louis, MO, USA
Niki Romani*, Innsbruck, Austria
Sem Saeland*, Lyon, France
Georg Stingl*, Vienna, Austria
Patrizia Stoitzner*, Innsbruck, Austria
Akira Takashima, Toledo, OH, USA
Kunihiko Tamaki*, Tokio, Japan
Past Workshops
1989 Oslo, Norway
1991 Lyon, France
1993 Dallas, TX, USA
1995 Scheveningen, The Netherlands
1997 Fuschl / Salzburg, Austria
1999 New York, USA
2001 Stresa, Italy
2003 Tokyo, Japan
2005 Funchal, Madeira, Portugal
2007 Berne, Switzerland
2009 Funchal, Madeira, Portugal
2011 Innsbruck, Austria
2 GENERAL INFORMATION
Participation fees
Early bird period
(payment received by August 1, 2013)
Payment
after August 1, 2013
Registration fee
425,- Euro
525,- Euro
Students*
300,- Euro
425,- Euro
*should provide proof of their student's or training status with a certificate of the University resp. Head
of Department
Registration hours
During the whole conference the registration desk in the foyer will be open 30 minutes before the daily
program starts and will close 15 minutes after the end of the daily program. Please contact us for any
question you may have on the meeting.
Congress Organization
Scientific contact & LC2013 website
Registration / Payment
Marcel Teunissen
Department of Dermatology
Academic Medical Center
Meibergdreef 9
1105 AZ Amsterdam
The Netherlands
Phone: 0031-20-566 4812
[email protected]
PDL Kongressmanagement
Andreas Hofer Strasse 6
A-6020 Innsbruck, Austria
Phone: 0043 512 567303
[email protected]
http://www.pdl.at
Congress Venue
Royal Tropical Institute
(Koninklijk Instituut voor de Tropen)
Mauritskade 63
1092 AD Amsterdam
Tel: 00-31-20 56 88 711
Fax: 00-31-20 66 84 579
www.kit.nl
3 SPONSORS
This meeting would not be possible without the sponsoring of the following partners from the industry,
as well as non-industrial organizations.
We are very grateful to :
Silver Sponsors
Celgene, Hillingdon, UK
CellGenix, Freiburg, Germany
Janssen-Cilag, Tilburg, The Netherlands
TissueGnostics, Vienna, Austria
www.celgene.com
www.cellgenix.com
www.janssennederland.nl
www.tissuegnostics.com
Bronze Sponsors
BioLegend, San Diego, CA, USA
eBioScience, Vienna, Austria
Genentech, South San Francisco, CA, USA
Greiner Bio-One, Alphen a/d Rijn, The Netherlands
Hyglos GmbH, Bernried, Germany
Lonza Pharma, Verviers,Belgium
Miltenyi Biotec, Leiden, The Netherlands
R & D Systems Europe Ltd, Abingdon, UK
www.biolegend.com
www.ebioscience.com
www.gene.com
www.greinerbioone.com
www.hyglos.de
www.lonza.com
www.miltenyibiotec.com
www.RnDSystems.com
Sponsors
Academic Medical Center, Amsterdam, The Netherlands
Celldex Therapeutics, Phillipsburg, USA
Dendritics, Lyon, France
ee-asi, a European Project (FP7 programme)
Stichting Nationaal Huidfonds, Utrecht, The Netherlands
IBL International, Amersfoort, The Netherlands
Leica Microsystems, Rijswijk, The Netherlands
LEO, Amsterdam, The Netherlands
Sanquin, Amsterdam, The Netherlands
Springer, Heidelberg, Germany
tebu-bio, Heerhugowaard, The Netherlands
The Rockefeller University Press, New York, USA
www.amc.nl
www.celldextherapeutics.com
www.dendritics.net
www.lc2013.nl
www.huidfonds.nl
www.IBL-International.com
www.leica-microsystems.com
www.leo-pharma.nl
www.sanquin.nl/en
www.springeronline.com
www.tebu-bio.com
www.rupress.org
Picture of the tulips on the front by courtesy of Tineke van der Pouw Kraan.
4 SPONSORS
Silver Sponsors
Bronze Sponsors
5 SPONSORS
Further Sponsors
6 LC2013 SYNOPSIS
Thursday
October 10, 2013
Friday
October 11, 2013
Saturday
October 12, 2013
Sunday
October 13, 2013
Registration
Registration
Registration
Session 1
Session 3
Session 5
Ontogeny &
Immunobiology
Tolerance &
Immunity
Vaccination
Coffee break
Coffee break
Coffee break
11:30
Session 1 continued
Session 3 continued
Session 5 continued
Vaccination
12:00
Ontogeny &
Immunobiology
Tolerance &
Immunity
08:30
09:00
09:30
10:00
10:30
11:00
12:30
13:00
Lunch &
Poster viewing
13:30
14:00
Lunch &
Poster viewing
Wrapping-up,
Awards, Invitations,
Concluding Remarks
Lunch
(only for attendees of
Satellite meeting)
14:30
Session 2
Session 4
Infection & Pattern
Recognition
Clinical Immunology
satellite meeting
by EE-ASI
consortium
Coffee break
Coffee break
"Antigen presentation
in the skin"
15:00
15:30
16:00
16:30
17:00
Session 2 continued
17:30
Infection & Pattern
Recognition
18:00
Registration
Session 4 continued
Clinical Immunology
18:30
19:00
19:30
Opening
Session
20:00
20:30
21:00
21:30
Social
Evening
Welcome
Reception
22:00
7 SCIENTIFIC PROGRAM
October 10, 2013
OPENING SESSION
Chair: Marcel Teunissen
19:00
Marcel Teunissen – Welcome and Opening of LC2013
19:15
Georg Stingl – Medical University of Vienna, Vienna, Austria
Langerhans cell research 1973 – 2013 : a journey into uncertainty
19:45
Frank Nestle – St. John's Institute of Dermatology, King's College, London, UK
Dermal dendritic cells: key sentinels of the skin immune system
20:30
Welcome Reception with Buffet
8 October 11, 2013
SCIENTIFIC PROGRAM
SESSION 1 — ONTOGENY & IMMUNOBIOLOGY
Chairs: Adelheid Elbe-Bürger & Kenji Kabashima
09:00
Florent Ginhoux – Singapore Immunology Network (SIgN), Singapore
Ontogeny of dendritic cells
09:30
Herbert Strobl – Medical University of Vienna, Vienna, Austria
Molecular mechanisms underyling Langerhans cell differentiation
10:00
Thomas Hieronymus – RWTH Aachen University, Aachen, Germany
Long-term and short-term Langerhans cell homeostasis
10:30
Coffee break
Chairs: Bianca Blom & Frank Nestle
11:00
Keisuke Nagao – Keio University School of Medicine, Tokyo, Japan
Hair follicles regulate immune homeostasis in skin
11:30
Barbara Schraml
Defining dendritic cells by ontogeny
11:45
Jia Tong Loh
The physiological role of polycomb group protein Ezh2 in Langerhans cells
12:00
Javed Mohammed
Keratinocytes control Langerhans cell homeostasis by TGFβ1 activating
Integrin β6
12:15
Benjamin Voisin
Homeostatic regulation of steady-state murine Langerhans cells under the
control of the hair follicle cycle
12:30
Thomas Bauer
Vitamin K-dependent receptors in control of Langerhans cell differentiation
and skin immunity
12:45
Kenji Kabashima
Inducible Skin-Associated Lymphoid Tissues (iSALT): An essential
architecture for efficient memory T cell activation in the skin
13:00
Lunch & Poster Viewing
October 11, 2013
SESSION 2 — INFECTIONS & PATTERN RECOGNITION
Chairs: Patrizia Stoitzner & Keisuke Nagao
14:30
James Young – Sloan-Kettering Institute for Cancer Research, New York, USA
Molecular clues to the superiority of human CD34+ HPC-derived
Langerhans cells for stimulating CTLs
15:00
Eynav Klechevsky – Washington University, St.Louis, MO, USA
What makes human Langerhans cells so special? Lessons learned using
systems biology approaches
15:30
16:00
Teunis Geijtenbeek – Academic Medical Center, University of Amsterdam, The
Netherlands
C-type lectins in infection and immunity
Coffee break
Chairs: Esther de Jong & Florent Ginhoux
16:30
Dan Kaplan – University of Minnesota, Minneapolis, MN, USA
Skin dendritic cell subsets and T helper cell phenotype
17:00
Sakeen Kashem
Candida albicans engagement of Dectin-1 on LC is required for IL-6
induction and Th17 differentiation
17:15
André Said
Autophagy restricts IL-23 secretion in monocyte-derived Langerhans-like
cells
17:30
Carla Ribeiro
Autophagy restricts HIV-1 infection in human Langerhans cells
17:45
Wilfried Posch
IgG opsonization impedes DC-mediated HIV-CTL priming
18:00
Yonatan Ganor
Calcitonin Gene-Relate Peptide (CGRP) inhibits LC-mediated HIV-1
transmission via an autocrine/paracrine feedback mechanism
18:15
Karin Loser
RANK-RANKL-activated Langerhans cells are critically involved in the upregulation of MHC class I-restricted anti-viral immunity during Herpes
simplex virus infections
18:30
Assembly of the International Committee on Langerhans Cell Workshops
10 October 12, 2013
SCIENTIFIC PROGRAM
SESSION 3 — TOLERANCE & IMMUNITY
Chairs: Eynav Klechevsky & Teunis Geijtenbeek
09:00
Michael Girardi – Yale University School of Medicine, New Haven, CT, USA
Unintended roles of Langerhans cells in cutaneous carcinogenesis
09:30
Muzlifah Haniffa – Newcastle University, Newcastle upon Tyne, UK
Human dendritic cell subsets
10:00
10:30
Esther de Jong – Academic Medical Center, University of Amsterdam, The
Netherlands
Functional specialization of skin dendritic cell subsets
Coffee break
Chairs: Muzlifah Haniffa & Georg Stingl
11:00
Knut Schäkel – University of Heidelberg, Germany
Slan (6-sulfo LacNAc) dendritic cells: a pro-inflammatory population of
dermal dendritic cells
11:30
Tal Capucha
Renewal of oral Langerhans cells in steady state and inflammatory
conditions
11:45
Sandrine Dubrac
The xenobiotic receptor pregnane X receptor modulates migration and
carcinogen metabolism in Langerhans cells
12:00
Carolina Martinez-Cingolani
Molecular mechanisms implicated in Thymic Stromal Lymphopoïetininduced dendritic cell migration
12:15
Thomas Döbel
Immune complex-mediated recruitment of slanDCs and NK cells to vascular
endothelium
12:30
Botond Igyártó
Langerhans cells and Langerin+ dermal dendritic cells promote humoral
responses through the generation of follicular helper T cells
12:45
Elodie Segura
Characterization of the intrinsic cross-presentation ability of human
dendritic cell subsets
13:00
Lunch & Poster Viewing
October 12, 2013
SESSION 4 — CLINICAL IMMUNOLOGY
Chairs: James Young & Matthew Collin
14:30
Bertrand Dubois – INSERM, Lyon, France
Langerhans cells dictate skin reactivity to weak contact allergens
15:00
Natalija Novak – University of Bonn Medical Center, Bonn, Germany
The role of human Langerhans cells of the oral mucosa and the skin in
allergic diseases
15:30
Björn Clausen – University Medical Center, Mainz, Germany
Dissecting the dendritic cell network in psoriasis-like skin disease
16:00
Coffee break
Chairs: Niki Romani & Björn Clausen
16:30
Matthew Collin – Newcastle University, Newcastle upon Tyne, UK
Langerin expression and mutated BRAF in human dendritic cells: more
clues about the origin of Langerhans cell histiocytosis
17:00
Elisabeth Glitzner
The role of dendritic cells in psoriasis
17:15
Rikke Bech
Interleukin 20 protein locates to distinct mononuclear cells in psoriatic skin
17:30
Rieneke van de Ven
Transcriptional profiling of human Langerhans cells in tumor draining
sentinel lymph nodes
17:45
Felix Scholz
Langerhans Cells suppress innate skin inflammation via liver CXCR6+ NK
cells
18:00
Merilyn Hibma
Langerhans cell homeostasis and activation is altered in epidermis
expressing human papillomavirus type 16 E7
18:15
Ilona J. Kosten
Technical advance: anti-CXCL12 inhibits mature LC migration in immunocompetent skin equivalents exposed to allergens
19:30
Social Evening with Dinner
12 October 13, 2013
SCIENTIFIC PROGRAM
SESSION 5 — VACCINATION
Chairs: Sandrine Henri & Dan Kaplan
09:00
09:30
Béhazine Combadière – INSERM, Paris, France
Langerhans cell's pivotal role in tailored immunity to vaccines
Yvette van Kooyk – Free University Medical Center, Amsterdam, The
Netherlands
Glycan-based targeting of vaccines to Langerhans cells and dendritic cells
in the skin
10:00
Laura Lozza
Crosstalk between human DC subsets promotes anti-mycobacterial activity
and CD8 T cell stimulation in response to tuberculosis vaccine Bacille
Calmette–Guérin
10:15
Marija Zaric
Efficient cross-priming and Th1 induction by Langerhans cells following
microneedle-mediated nanoparticle transdermal vaccination
10:30
Coffee break
Chairs: Yvette van Kooyk & Knut Schäkel
11:00
Adrien Kissenpfennig – Queens University Belfast, Belfast, Uk
Can Langerhans cells take the sting out of vaccines?
11:30
Karsten Mahnke
Targeting of myelin oligodendrocyte glycoprotein to the DC/LC antigen
receptor DEC205 in vivo induces immune suppression and prevents allergic
experimental encephalomyelitis (EAE) in mice
11:45
Cynthia M. Fehres
C-type lectin mediated uptake of glycan modified antigens by primary
human Langerhans cells results in cross presentation to CD8+ T cells
12:00
Niki Romani – Innsbruck Medical University, Innsbruck, Austria
Wrapping up LC2013: the current status of Langerhans cells, their future
and beyond
12:30
Huidfonds Travel Award & Rockefeller Press Travel Award
Greiner Poster Award & Springer Poster Award
12:45
Elodie Segura – Invitation to DC2014 in Tours, France
12:50
Keisuke Nagao, Tatsuyoshi Kawamura and Kenji Kabashima
– Invitation to LC2015 in Kyoto, Japan
12:55
Concluding remarks and Closing LC2013
13 SATELLITE MEETING
October 13, 2013
EE-ASI FP7 programme – LC2013 satellite meeting
“Antigen presentation in the skin”
Agenda
Morning: Joint session with the LC2013 (Vaccination) 13.00 – 14.00 Lunch (registration) Afternoon: 1st EE‐ASI Workshop 14.00 – 14.05 Welcome – Sandrine Henri, Inserm, Marseille, France 14.05 – 14.15 The EE‐ASI programme – Colin Dayan, Cardiff University 14.15 – 14.45 Patrizia Stoitzner, Innsbruck Medical University, Innsbruck, Austria: “Combination of molecular targeted therapy and immunotherapy: the future of personalized medicine” 14.45 – 15.15 Marc Vocanson, University of Lyon, Lyon, France: "Skin tolerance to haptens" 15.15 – 15.45 2 Short oral presentations 15.45 – 16.15 Coffee break 16.15 – 16.45 Yasmine Belkaid, National Institutes of Health, Bethesda, USA: “Microbiota: The skin secret weapon.” 16.45– 17.15 Michael Sixt, Institute of Science and Technology Austria, Klosterneuburg, Austria: “From the skin to the LN” 17.15 – 17.30 Concluding remarks The short oral presentations of the afternoon session will be selected from abstracts submitted to the EE‐ASI Workshop and to be addressed to Sandrine HENRI ([email protected]‐mrs.fr) and Marie Clotteau (anna.boitard@inserm‐transfert.fr). EE‐ASI is a collaborative research project supported by the Health Cooperation work programme of the 7th Framework Programme for the Research and Technological Development of the European Commission.
15 INVITED SPEAKER
October 10, 2013
Georg Stingl – Medical University of Vienna, Vienna, Austria
Langerhans cell research 1973 – 2013 : a journey into uncertainty
Space for your notes :
16 19.15 – 19.45
19.45 – 20.15
October 10, 2013
INVITED SPEAKER
Frank Nestle – St. John's Institute of Dermatology, King's College, London, UK
Dermal dendritic cells: key sentinels of the skin immune system
17 INVITED SPEAKER
October 11, 2013
Florent Ginhoux – Singapore Immunology Network (SIgN), Singapore
Ontogeny of dendritic cells
18 09.00 – 09.30
09.30 – 10.00
October 11, 2013
Herbert Strobl – Medical University of Vienna, Vienna, Austria
Molecular mechanisms underyling Langerhans cell differentiation
19 INVITED SPEAKER
INVITED SPEAKER
October 11, 2013
Thomas Hieronymus – RWTH Aachen University, Aachen, Germany
Long-term and short-term Langerhans cell homeostasis
20 10.00 – 10.30
11.00 – 11.30
October 11, 2013
Keisuke Nagao – Keio University School of Medicine, Tokyo, Japan
Hair follicles regulate immune homeostasis in skin
21 INVITED SPEAKER
SELECTED ABSTRACT
October 11, 2013
11.30 – 11.45
Defining dendritic cells by ontogeny
Barbara U. Schraml1, Janneke van Blijswijk1, Santiago Zelenay1, Paul G. Whitney1, Andrew Filby2, Neil
C. Rogers1, Caetano Reis e Sousa1
1
Immunobiology Laboratory, , Cancer Research UK, London Research Institute, Lincoln's Inn Fields
Laboratories, London WC2A 3LY, United Kingdom
2
Flow Cytometry Laboratory, Cancer Research UK, London Research Institute, Lincoln's Inn Fields
Laboratories, London WC2A 3LY, United Kingdom
Mononuclear phagocytes sample the environment for signs of damage or infection. The classification
of these cells as macrophages or dendritic cells (DC) has traditionally been done on the basis of
differences in cell morphology, expression of specific markers or of select functional attributes.
However, these attributes are not absolute and often overlap, leading to difficulties in cell type
identification. To circumvent these issues, we have generated a model to define DC based on their
ontogenetic descendence from a committed precursor.
We show that in mice precursors of conventional DC but not other leukocytes are marked by
expression of DNGR-1/CLEC9A. We generated a mouse model to genetically label Clec9a-expressing
conventional DC precursors and their progeny with yellow fluorescent protein (YFP). Genetic labeling
of these cells and their progeny specifically traces cells traditionally ascribed to the DC lineage and the
restriction is maintained after infection. Notably, in some tissues cells previously thought
monocytes/macrophages are in fact descendants from DC precursors. These studies provide the first
in vivo model for lineage tracing of DC and allow the definition of DC based on ontogenetic rather than
phenotypic, morphological or functional criteria. These studies establish DC as an independent
immune lineage and distinguish them from other leukocytes, thus paving the way to unraveling the
functional complexity of the mononuclear phagocyte system.
Notes:
22 11.45 – 12.00
October 11, 2013
SELECTED ABSTRACT
The physiological role of polycomb group protein Ezh2 in Langerhans cells
Jia Tong Loh 1, Merry Gunawan 1, Florent Ginhoux 2, I-Hsin Su 1
1
Division of Molecular Genetics and Cell Biology, School of Biological Sciences, College of Science,
Nanyang Technological University, Singapore
2
Singapore Immunology Network, Agency for Science, Technology and Research, Singapore
Polycomb group protein Enhancer of Zeste homolog 2 (Ezh2) is a histone methyltransferase which
regulates gene expression through the modification of chromatin structure. Our recent studies have
demonstrated that Ezh2 can act on a novel cytosolic substrate talin in dendritic cells (DCs), thereby
regulating DCs’ integrin signaling and adhesion dynamics. Here, we have identified the expression of
Ezh2 to be pivotal in the regulation of Langerhans cells (LCs) migration during steady state and under
inflammatory conditions. Even though Ezh2-deficient LCs were able to disengage from the
surrounding keratinocytes, they failed to exit from the epidermis, leading to their reduced frequencies
observed in the dermis and skin-draining lymph nodes. In particular, our data highlighted a crucial role
of Ezh2 in mediating LCs transmigration across the basement membrane, as the migratory capacity of
Ezh2-deficient LCs became attenuated upon encountering this barrier situated between the epidermis
and dermis. These LCs were observed to form enlarged focal adhesions, which led to their enhanced
spreading on laminin, a major constituent of the basement membrane. Hence, Ezh2 may regulate skin
immunity through its control on LCs migration. The molecular mechanisms regulating these processes
will be further determined.
Notes:
23 SELECTED ABSTRACT
October 11, 2013
12.00 – 12.15
Keratinocytes control Langerhans cell homeostasis by TGFβ1 activating Integrin β6
Javed Mohammed, Elizabeth S. Jarrett, Sakeen Kashem, Daniel H. Kaplan
Department of Dermatology, Center for Immunology, University of Minnesota, Minneapolis, MN 55455,
USA
Ablation of Transforming Growth Factor Beta1 (TGFβ1) or the TGFβ type II receptor selectively in
Langerhans cells (LC) leads to their spontaneous migration to skin-draining lymph nodes (LN). TGFβ1
is secreted as a homodimer that is non-covalently complexed with the latency-associated-peptide
(LAP). TGFβ1 activators mediate LAP/TGFβ1 dissociations thereby allowing active TGFβ1 homodimer
to bind to its receptor. Mice with a mutated RGD sequence in LAP completely lack epidermal LC
suggesting a requirement of RGD-binding integrin αvβ6 (itgβ6) for epidermal LC homeostasis. αvβ6 is
expressed by bulge keratinocytes (KC) and interfollicular epidermal (IFE) KC subsets but is absent
from hematopoietic cells within the epidermal compartment. Since bulge KC inhibit recruitment of LC
precursors in to the epidermis, we hypothesized that αvβ6 expressed by IFE KC is required to prevent
spontaneous LC migration. Consistent with our hypothesis, adult Itgβ6-/- mice have fewer LC that
were distributed near hair follicles but were absent from the IFE. There were also increased numbers
of total LC and BrdU+ LC in the LN of itgβ6-/- mice compared with controls. In addition, a higher
percentage of epidermal LC in Itgβ6-/- mice expressed Ki67 and intradermal injection of Itgβ6
neutralizing antibody resulted in LC activation. To confirm that KC derived Itgβ6 is required for
epidermal retention of steady-state LC, we generated bone marrow chimeras with WT or Itgβ6-/- mice
as recipients of congenic WT bone marrow cells. WT LC failed to populate epidermis of Itgβ6-/recipients and had similar distribution to Itgβ6-/- recipient LC demonstrating that KC-derived Itgβ6 is
required to maintain LC homeostasis. We also observed that IFE reduce expression of αvβ6 in
response to exposure to UV light. These data are consistent with a model in which regulated
expression of αvβ6 by KC can control LC activation and migration.
Notes:
24 12.15 – 12.30
October 11, 2013
SELECTED ABSTRACT
Homeostatic regulation of steady-state murine Langerhans cells under the control of the hair
follicle cycle
Benjamin Voisin1, Dimitri Chartoire1, Pascal Schneider2, Christine Kowalczyk2, Vincent Flacher1 and
Christopher G. Mueller1
1
CNRS UPR3572 Immunopathologie et Chimie Thérapeutique, Institut de Biologie Moléculaire et
cellulaire, Strasbourg, France.
2
Department of Biochemistry, University of Lausanne, Epalinges, Switzerland
Hair follicle (HF) is a skin dynamic structure which, after establishing at embryonic stage
(morphogenesis), undergoes continuous renewing in a cycle composed of three phases: growth
(anagen), apoptotic regression (catagen) and resting (telogen). This cycle triggers HF microenvironment modifications. Langerhans cells (LCs) form a dense epithelial network, maintained by an
in situ LC self-renewal, and show close association with HFs. In juvenile wild-type mice, which display
synchronized HF cycle, we observed that LCs had a higher rate of proliferation during the anagen.
Anagen-associated enhancement of LC renewal occurred mostly around HFs and was independent of
a CX3CR1+ LC precursor infiltration. Accordingly, inducing anagen by depilation of wild-type adult
mice led to increased proliferation of LCs. On the other hand, upon interruption of HF cycle in the
Rankl-/- mice, LCs displayed strongly reduced proliferation ability. Moreover, in Eddar-/- mice lacking
HF morphogenesis in the tail skin, LCs did establish an epidermal network but their self-renewal was
not promoted in anagen. Finally, in addition to proliferative bursts, HF cycle phases correlated with LC
migration waves, since lymph nodes draining catagen/telogen skin contained more LCs than those
draining anagen skin. Altogether, our results bring further insight into LC homeostasis, showing that
the HF exerts a critical control on LC proliferation and emigration in steady-state mouse skin
throughout life.
Notes:
October 11, 2013
12.30 – 12.45
Vitamin K-dependent receptors in control of Langerhans cell differentiation and skin immunity
Thomas Bauer1,2, Anna Zagórska3, Jennifer Jurkin1, Nighat Yasmin1, René Köffel1,2, Susanne Richter1,
Bernhard Gesslbauer1, Greg Lemke3,4 and Herbert Strobl1,2
1
Institute of Immunology, Center of Pathophysiology, Infectiology and Immunology, Medical University
of Vienna, 1090 Vienna, Austria
2
Institute of Pathophysiology and Immunology, Center of Molecular Medicine, Medical University
Graz, 8036 Graz, Austria
3
Molecular Neurobiology Laboratory
4
Immunobiology and Microbial Pathogenesis Laboratory, The Salk Institute, La Jolla, CA 92037
On the outermost edge of the body a dense network of dendritic cells (DCs), the Langerhans cells
(LCs), represent the first immune barrier. The establishment and maintenance of this epidermal
network is dependent on the cytokine transforming growth factor-beta1 (TGF-β1) expressed by
keratinocytes (KC) and LCs. We could recently identify a crucial downstream effector of TGF-β1, the
receptor tyrosine kinase Axl. Axl belongs to the TAM receptor family, with the additional members
Tyro3 and Mer, and gets activated through the vitamin K-dependent ligands Gas6 and Protein S. Axl
mediates parts of the TGF-β1 effects on LC differentiation, enhancement of phagocytosis and
inhibition of pro-inflammatory cytokine signalling. Moreover, the activity of the TAM receptors was
crucial for resolving antigen-specific T-cell mediated skin inflammation in mice. In line with this,
constitutive Axl is activated and Mer is neo-induced in human skin LCs in response to allergens and
both of these genes are controlled by TGF-β1. Dysfunction of these receptors might lead to
autoimmune/ inflammatory skin diseases such as lupus erythematosus, psoriasis or atopic dermatitis.
We now aim to better characterize downstream effectors of TAM receptors during LC differentiation/
activation. Addition of vitamin K to serum-free human LC differentiation cultures, in order to activate
the endogenous ligands Gas6 and Protein S, results in Axl dependent positive effects on LC
differentiation and the expression of several LC specific and anti-inflammatory signature genes.
Additionally, we identified for the first time a second vitamin K-dependent receptor-ligand system
downstream of TGF-β1 in LCs, the Protein C receptor system, which is implicated in wound healing
and DC specific anti-inflammatory effects. Together, these receptors might secure immunologic
tolerance at body surfaces.
Notes:
26 12.45 – 13.00
October 11, 2013
SELECTED ABSTRACT
Inducible Skin-Associated Lymphoid Tissues (iSALT): An essential architecture for efficient
memory T cell activation in the skin
Kenji Kabashima
Department of Dermatology, Kyoto University Graduate School of Medicine
Although a key step during secondary challenge is antigen presentation to peripheral memory T cells,
it remains largely unclear how antigen presenting cells and memory T cells encounter each other
efficiently in the periphery. In the skin, the concept of skin-associated lymphoid tissues (SALT) has
been proposed; however, what happens to various components of SALT following antigen exposure is
not well characterized. Here, we report the identification of immune cell clusters in the skin induced by
antigen challenge, which we call inducible SALT (iSALT), using a murine contact hypersensitivity
(CHS) model. We report that dermal dendritic cells (DCs), but not epidermal Langerhans cells, were
required for the elicitation of CHS and that DCs formed clusters in dermal perivascular areas and that
they interacted with skin-infiltrating memory T cells for several hours. This sustained interaction was
required for in situ proliferation and activation of memory T cells in an antigen- and integrin LFA-1dependent manner. Intriguingly, DC clustering in the dermis was abrogated by depletion of skin
macrophages, and DCs were attracted toward macrophages both in vivo and in vitro. Furthermore, IL1α treatment induced CXCL2 production from M2-phenotype macrophages, and DC clustering was
suppressed with blockade of either IL-1R or CXCR2. Taken together, our findings suggest that dermal
DCs and memory T cells form immune cell clusters, iSALT, for their efficient encounter in the skin via
macrophage activation by IL-1. This sustained conjugation between DCs and memory T cells is
essential for establishment of the effector phase of acquired cutaneous immunity.
Notes:
27 INVITED SPEAKER
October 11, 2013
14.30 – 15.00
James Young – Sloan-Kettering Institute for Cancer Research, New York, USA
Molecular clues to the superiority of human CD34+ HPC-derived Langerhans cells for
stimulating CTLs
28 15.00 – 15.30
October 11, 2013
INVITED SPEAKER
Eynav Klechevsky – Washington University, St.Louis, MO, USA
What makes human Langerhans cells so special? Lessons learned using systems
biology approaches
29 INVITED SPEAKER
October 11, 2013
15.30 – 16.00
Teunis Geijtenbeek – Academic Medical Center, University of Amsterdam, The Netherlands
C-type lectins in infection and immunity
30 16.30 – 17.00
October 11, 2013
Dan Kaplan – University of Minnesota, Minneapolis, MN, USA
Skin dendritic cell subsets and T helper cell phenotype
31 INVITED SPEAKER
SELECTED ABSTRACT
October 11, 2013
17.00 – 17.15
Candida albicans engagement of Dectin-1 on LC is required for IL-6 induction and Th17
differentiation
Sakeen W. Kashem, Botond Z. Igyártó, Daniel H. Kaplan
Department of Dermatology, Center for Immunology, University of Minnesota, MN
Infection with extracellular bacteria and fungi are a global burden resulting in millions of
hospitalizations and deaths per year. T helper 17 (Th17) cells are a recently appreciated subset of
CD4+ helper cells that are involved in protection against such pathogens. During a superficial skin
infection with Candida albicans we have previously found that Langerhans cells (LC) and CD103+
dermal DC (dDC) were necessary and sufficient for in vivo induction of Th17 and Th1, respectively. To
explore whether CD11b+ dDC participate in induction of Th17, we infected huLangerin-DTA x BatF3-/mice with lack both LC and CD103+ dDC, as well as MGL-2DTR-eGFP/+ mice that allow for the DT
induced ablation of most CD11b+ dDC. In mice lacking LC and CD103+ dDC, T cell proliferation was
unaffected but induction of both Th1 and Th17 was defective. In contrast, in the absence of most
CD11b+ dDC, T cell proliferation was reduced but induction of Th1 and Th17 was unaffected. Th17
differentiation depends on the cytokines IL-1β, IL-6, TGF-β and IL-23 that are all expressed by LC. We
generated mice with LC-specific ablation of each cytokine using either cre-lox or bone-marrow
chimeras. Only LC-derived IL-6 was required for Th17 induction. Notably, we found that skin infection
with a strain of C. albicans that does not engage Dectin-1 resulted in diminished LC expression of IL-6
and failed to induce Th17 differentiation. Thus, we have identified that each of the 3 major skin DC
subsets has a unique and non-redundant function during adaptive responses to C. albicans skin
infection: CD11b+ dDC are required for optimal T cell proliferation; CD103+ dDC are required for Th1;
and LC via Dectin-1 mediated induction of IL-6 are required for Th17.
Notes:
32 17.15 – 17.30
October 11, 2013
SELECTED ABSTRACT
Autophagy restricts IL-23 secretion in monocyte-derived Langerhans-like cells
André Said, Günther Weindl
Institute of Pharmacy (Pharmacology and Toxicology), Freie Universität Berlin, Berlin, Germany
Recent studies suggest a role for autophagy in the secretion of IL-1 cytokines, thereby regulating and
affecting the development of inflammatory diseases. The antimalarial drug and autophagy inhibitor
chloroquine has long been considered as a potential trigger of drug-induced or drug-aggravated
psoriasis, where mainly activated Th17 cells sustain a persistent inflammation. Here, we investigated
the effect of IL-1beta on monocyte-derived Langerhans-like cells (MoLC) and dendritic cells (MoDC) in
the presence of chloroquine as a lysosomotropic agent. To assess potential alterations in the release
of Th1 or Th17 related cytokines, we stimulated MoLC and MoDC with IL-1beta alone or in
combination with TNF and different TLR ligands. Interestingly, treatment with chloroquine enhanced
IL-6 production but reduced IL-12p70 release in both subsets and IL-23 in MoDC, whereas MoLC
showed strongly increased IL-23 secretion. Moreover, intracellular FACS analysis revealed a decrease
in IL12p40 production, confirming the lower IL-12p70 release in both subsets. The enhanced IL-23
release in response to chloroquine was dependent on IL-1beta and IL-1alpha and also observed with
the autophagy inhibitors hydroxychloroquine and bafilomycin. Treatment with the specific TGF-betaR1
inhibitor LY364947 did not suppress IL-23 release by activated MoLC, excluding a critical involvement
of TGF-beta as key regulator of MoLC differentiation. Noteworthy, despite the similarity in IL-1R and
TLR2/MyD88 signaling, stimulation with the TLR2/1 ligand Pam3CSK4 in the presence of chloroquine
induced IL-23 in both MoLC and MoDC. Taken together, our findings suggest a potentially essential
role of human Langerhans cells in chloroquine-provoked psoriasis through promotion of Th17
differentiation by increasing IL-23 and IL-6 levels in respective skin lesions.
Notes:
October 11, 2013
17.30 – 17.45
Autophagy restricts HIV-1 infection in human Langerhans cells
Carla M. S. Ribeiro, Ramin Sarrami-Forooshani, Linda M. van den Berg, Sonja I. Gringhuis and Teunis
B.H. Geijtenbeek
Department of Experimental Immunology, Academic Medical Center, University of Amsterdam,
Sexual transmission across genital mucosa is the main route of HIV-1 infection. Langerhans cells
(LCs) reside in the mucosal epithelia such as vagina, foreskin as well as in the skin epidermis and are
the first immune cells to encounter HIV-1 upon sexual transmission. LCs express the C-type lectin
receptor Langerin which prevents HIV-1 infection of LCs and subsequent HIV-1 transmission. Langerin
captures HIV-1 into Birbeck granules where the virus is thought to be degraded. Although this
Langerin-specific internalization pathway is central to the ability of LCs to degrade HIV-1, the function
of Birbeck granules and the molecular mechanisms involved in HIV-1 degradation are largely
unknown. Autophagy can function as an antiviral mechanism by targeting viral components or virions
for lysosomal degradation. Here we investigated the role of autophagy during HIV-1 infection in LCs.
The molecular mechanisms involved in viral clearance by LCs were investigated by using inducers or
inhibitors of the autophagy pathway and by silencing key components of the autophagy pathway. HIV1 induces autophagy in LCs and boosting autophagy-mediated lysis enhances HIV-1 degradation in
LCs. Moreover, loss of autophagy-mediated lysis by silencing autophagosomal molecules Atg5 and
Atg16L1 resulted in a striking increase of viral DNA integration into the host genome in LCs. Coimmunoprecipitation studies confirmed the co-localization of Langerin and autophagosomal molecules
after HIV-1 internalization in LCs. This suggests a protective role for autophagy in HIV-1 infection after
virus uptake. In addition, our findings that Birbeck granules are caveolar vesicles further suggests that
caveolar-mediated uptake is linked to viral degradation. Our data show that the LC-specific
internalization route for HIV-1 intersects with the autophagy pathway and that the autophagy pathway
is essential to mount an effective antiviral immune response against HIV-1 in LCs.
Notes:
34 17.45 – 18.00
October 11, 2013
SELECTED ABSTRACT
IgG opsonization impedes DC-mediated HIV-CTL priming
Wilfried Posch1, Gianfranco Pancino2, Cornelia Lass-Flörl1, Arnaud Moris3, Asier Saez-Cirion2, Doris
Wilflingseder1
1
Division of Hygiene and Medical Microbiology, Innsbruck Medical University, Schöpfstrasse 41/311,
Innsbruck, Austria
2
Unité de Régulation des Infections Rétrovirales, Institut Pasteur, 25 Rue du Docteur Roux, 75724
Paris, France
3
INSERM UMRS-945, Infection and Immunity, Hôpital La Pitié-Salpêtrière, Université Pierre et Marie
Curie (Paris-6), 91 Bd de l´Hôpital, Paris, France
Potent effector functions of the HIV-specific CD8+ T cell (CTL) response are supposed to be
responsible to control the virus. The opsonization pattern of the pathogen can modify antigen capture,
its presentation as well as the priming of specific CTL responses. Based on our observation that
higher levels of antibodies are deposited on the HIV surface using plasma from HIV-positive
individuals with higher viral loads, we analyzed the impact of IgG-opsonization on the antigenpresenting capacity of DCs. We here show that IgG opsonization of the virus is associated with a loss
of the CTL-stimulatory capacity by DCs as represented by reduced proliferation, low activation of HIVspecific CTL clones and a weak antiviral activity. Our ex vivo and in vitro observations illustrate a close
correlation between the HIV opsonization pattern and DC-induced expansion and differentiation of
specific CTLs. They are relevant regarding HIV vaccination strategies as they suggest that strong,
transient antibody responses after vaccination might contribute to weaken CTL-induction by
modulation of DC function.
Notes:
October 11, 2013
18.00 – 18.15
Calcitonin Gene-Relate Peptide (CGRP) inhibits LC-mediated HIV-1 transmission via an
autocrine/paracrine feedback mechanism
Yonatan Ganor1,2,3, Anne-Sophie Drillet-Dangeard1,2,3, Christian Federici1,2,3, Morgane Bomsel1,2,3
1
Mucosal Entry of HIV-1 and Mucosal Immunity, Department of Infection, Immunity and Inflammation,
Cochin Institute, CNRS (UMR 8104), Paris, France
2
INSERM U1016, Paris, France
3
Paris Descartes University, Sorbonne Paris Cité, Paris, France
In mucosal epithelia, peripheral neurons are in contact with resident LCs. We recently reported
(Ganor, JEM, in press) that the peripheral neuropeptide Calcitonin Gene-Related Peptide (CGRP)
strongly inhibits mucosal HIV-1 transmission by interfering with multiple steps of LC-mediated HIV-1
transfer to T-cells. We used now differential gene arrays comparing untreated/CGRP-treated LCs and
substantiating experiments (RT-PCR, flow cytometry, fluorescent microscopy, ELISA) to further
understand the anti-HIV-1 potential of CGRP. This analysis confirms our previous findings that CGRP
activates NFκB, increases langerin and decreases integrins expression. Moreover, expression of the
HIV-1 co-receptor CCR5 appears decreased, while CXCR4 is increased, correlating with inhibition of
R5 but not X4 HIV-1 transfer by CGRP. New candidate proteins that might modulate HIV-1 binding,
internalization and degradation in LCs are also identified. Unexpectedly, transcription and expression
of CGRP itself and its cognate receptor in LCs are upregulated by CGRP. Hence, CGRP might
engage a positive feedback mechanism that would further enhance its anti-HIV-1 activity. Together,
these experiments demonstrate the complexity of the interactions between CGRP, LCs and HIV-1.
This information might be used therapeutically to enhance the anti-HIV-1 activity of CGRP. Finally, in
addition to their morphological resemblance with neurons, LCs present some actual neuronal
characteristics.
Notes:
36 18.15 – 18.30
October 11, 2013
SELECTED ABSTRACT
RANK-RANKL-activated Langerhans cells are critically involved in the up-regulation of MHC
class I-restricted anti-viral immunity during Herpes simplex virus infections
Lars Klenner1, Wali Hafezi2, Björn E. Clausen3, Thomas A. Luger1, Joachim E. Kühn2, Karin Loser1
1
Department of Dermatology, University of Münster, 48149 Münster, Germany
Medical Microbiology, University of Münster, 48149 Münster, Germany
3
Erasmus MC, University Medical Center, Department of Immunology, Rotterdam, The Netherlands
2
Infectious diseases frequently occur in humans and Herpes simplex virus type 1 (HSV-1) represents
the viral pathogen causing the majority of cutaneous infections. Viral infections are controlled by the
immune system and cytotoxic CD8+ effector T cells have been shown to play a crucial role during the
clearance of acute viral infections from the skin. In mouse models of cutaneous HSV-1 infection it has
been shown that effector T cell priming requires the uptake of HSV-1 antigens by epidermal
Langerhans cells (LC) and the subsequent antigen transfer to lymph node resident dendritic cells in a
CD86 dependent process. Since RANK-RANKL signalling is critical for dendritic cell – T cell
communication we investigated whether RANK-RANKL interactions might play a role in cutaneous
anti-viral immunity. Therefore, transgenic mice over-expressing RANKL (CD254) in basal
keratinocytes (K14-RANKL tg) were epicutaneously infected with HSV-1. Interestingly, K14 RANKL tg
mice developed significantly smaller skin lesions compared to wildtype (wt) controls, which was
accompanied by reduced numbers of virus particles and decreased virus replication in tg skin, thus
indicating increased primary anti-viral immune responses in K14 RANKL tg mice. In support of this,
flow cytometry and immunohistology revealed up-regulated levels of total CD8+ T cells as well as an
enhanced expression of markers associated with cytotoxic activity, like IFN-gamma, Fas ligand,
granzymes, perforin or KLRG1, in CD8+ T cells from K14-RANKL tg compared to wt mice. To analyze
the specific role of CD8+ T cells for anti-viral immune responses we isolated CD8+ effector T cells
from HSV-1-infected wt as well as K14-RANKL tg donors, transferred them into naive congenic
recipients and subsequently, infected recipient mice with HSV-1. The transfer of CD8+ T cells from
HSV-1-infected wt donors resulted in a slight reduction of skin lesion size compared to recipients that
received PBS instead of CD8+ T cells. However, mice that were injected with CD8+ T cells from HSV1-infected K14-RANKL tg donors failed to develop inflammatory skin lesions upon HSV-1 challenge
suggesting a strong MHC class I-restricted anti-viral immune response. To investigate whether CD8+
T cells are indeed essential for the increased anti-viral immunity in K14-RANKL tg mice we depleted
CD8+ T cells in wt and tg mice prior to HSV-1 infection using specific antibodies. Upon HSV-1
challenge CD8-depleted wt mice developed skin lesions similar to those observed in mice treated with
an IgG control antibody. Strikingly, CD8-depletion in K14-RANKL tg mice led to a substantial increase
in skin lesion size. Since during the induction of cutaneous anti-viral immunity LC represent the
primary DC subset getting into contact with the virus, we analyzed the effect of RANK-RANKL
signalling on the phenotype and function of LC. Whereas HSV-1 infection is known to induce
apoptosis in LC from wt skin TUNEL staining of lesional skin revealed that tg LC were protected from
virus-induced apoptosis. Moreover, we observed an up-regulated expression of the activation marker
CD86 in LC from HSV-1-infected tg skin potentially suggesting an increased T cell stimulatory
capacity. To elucidate the role of RANK-RANKL signaling on LC function during cutaneous HSV-1
infection in more detail K14-RANKL tg mice were bred to Langerin-DTR mice and LC were depleted
from double mutants by injecting diphtheria toxin prior to epicutaneous HSV-1 challenge. Notably, LCdepleted K14-RANKL tg mice developed skin lesions similar to wt controls indicating a critical role of
LC for the induction of anti-viral immunity in tg mice. Additionally, the depletion of LC from K14-RANKL
tg skin impaired the expansion of anti-viral CD8+ effector T cells. Together, these data indicate that
cutaneous RANK-RANKL signaling is critically involved in the priming of HSV-1-specific CD8+ effector
T cells. This effect was initiated by the up-regulation of activating co-stimulatory molecules as well as
the prevention of virus-induced apoptosis in LC.
Notes:
37 INVITED SPEAKER
October 12, 2013
Michael Girardi – Yale University School of Medicine, New Haven, CT, USA
Unintended roles of Langerhans cells in cutaneous carcinogenesis
38 09.00 – 09.30
09.30 – 10.00
October 12, 2013
Muzlifah Haniffa – Newcastle University, Newcastle upon Tyne, UK
Human dendritic cell subsets
39 INVITED SPEAKER
INVITED SPEAKER
October 12, 2013
10.00 – 10.30
Esther de Jong – Academic Medical Center, University of Amsterdam, The Netherlands
Functional specialization of skin dendritic cell subsets
40 11.00 – 11.30
October 12, 2013
INVITED SPEAKER
Knut Schäkel – University of Heidelberg, Germany
Slan (6-sulfo LacNAc) dendritic cells: a pro-inflammatory population of dermal
dendritic cells
October 12, 2013
11.30 – 11.45
Renewal of oral Langerhans cells in steady state and inflammatory conditions
Tal Capucha1, Gabriel Mizraji1, Hadas Segev1, Yaffa Shaul1, Katya Zelentsova1, Luba Eli-Berchoer1,
Asaf Wilensky2 and Avi-Hai Hovav1
1
Institute of Dental Sciences, Hebrew University-Hadassah Faculty of Dental Medicine, Jerusalem,
Israel
2
Department of Periodontology, Hebrew University-Hadassah Faculty of Dental Medicine, Jerusalem,
Israel
Whereas our understanding regarding the origin and function of epidermal Langerhans cells (LCs)
increased tremendously over the past decades, less is known about LCs located in the oral epithelium
(oLCs). As the most peripheral antigen presenting cells of the oral cavity, oLCs are constantly exposed
to the immense oral microbiota and are required to distinguish between innocuous and harmful
microbes. We recently demonstrated the critical role of oLCs in down-modulating local inflammation
and tissue destruction by oral pathogen. Given the importance of oLCs in oral immunity, it is crucial to
understand how these cells are maintained during steady-state condition and inflammation. By using
bone marrow chimeras we found that in contrast to epidermal LCs, oLCs are renewing from bone
marrow precursors. Continuous administration of BrdU indicated that oLCs are labeled in a faster
kinetics than epidermal LCs, where complete labeling was observed three weeks after treatment.
Similar renewing kinetics were observed following in vivo ablation of langerin-expressing cells in
langerin-DTR mice. To monitor the repopulation kinetics of oLCs under inflammatory condition, we
infected mice with the oral pathogen Porphyromonas gingivalis via an oral gavage. Two days after
infection, the frequencies of oLCs were considerably reduced in correlation with their arrival to the
cervical LNs. A gradual repopulation was observed on the following days whereas 10 days after
infection the frequencies of oLCs in the gingiva were completely restored. Taken together, we
demonstrated that in contrast to epidermal LCs, oral mucosal LCs originates from circulating bone
marrow precursors and are completely renewed within three weeks. Under inflammatory condition,
steady state oLCs departure the epithelium rapidly and the tissue is swiftly repopulated by
inflammatory LCs. Such differential ontogeny might also represent a divergent functional specialization
between oral and epidermal LCs.
Notes:
42 11.45 – 12.00
October 12, 2013
SELECTED ABSTRACT
The xenobiotic receptor pregnane X receptor modulates migration and carcinogen metabolism
in Langerhans cells
Matthias Schmuth1, Florian Sparber1, Andreas Elentner1, Susanne Ebner2, Barbara Del Frari3, and
Sandrine Dubrac1
1
Department of Dermatology and Venereology, Innsbruck Medical University, 6020 Innsbruck,
Austria.
2
VTT Daniel Swarovski Labor, Department of Surgery, Innsbruck Medical University, 6020 Innsbruck,
Austria
3
Department of Plastic, Reconstructive and Esthetic Surgery, Innsbruck Medical University, 6020
Innsbruck, Austria
Skin exposure to haptens, microbes or chemicals including carcinogens induces migration of skin DC.
The pregnane X receptor (PXR) is a transcription factor activated by xenobiotics, hormones and
cholesterol metabolites. We here found that PXR is expressed in different subsets of mouse and
human immature dendritic cells (DC) especially Langerhans cells (LC) where it controls CCR7
expression. Furthermore, PXR modulates expression of TGF-β, SMAD3 and SOCS-1 in epidermal
cells with expression of TGF-β preceding that of SOCS-1 and SMAD3. In LC, PXR signals via the
TGF-β pathway by preventing the down-regulation of active form of smad2/3 that normally occurs
upon LC maturation. Furthermore, neutralization of TGF-β reverses the effects of PXR activation on
CCR7, SOCS-1 and SMAD3 expression. In vivo, PXR deficiency increases migration of LC after skin
exposure to a hapten (TNCB) but also to a carcinogen (DMBA) and delays appearance of chemicallyinduced papilloma and squamous cell carcinoma. Promotion of LC migration in PXR deficient mice
after topical application of DMBA may result from a protection against DMBA-induced cell death that
occurs in wild type LC. Thus, PXR could be important for the metabolism of chemicals by LC.
Accordingly, we found that a single topical application of DMBA up-regulates PXR and its well-known
down-stream gene Cyp3a11 in the skin and that PXR deficiency protects skin against DNA damage
after a single topical application of DMBA. Together, these results demonstrate that PXR controls
CCR7 and LC migration and that PXR signals via TGF-β pathway. Moreover, PXR might be involved
in the metabolism of chemicals by LC and thus in cutaneous carcinogenesis.
Notes:
October 12, 2013
12.00 – 12.15
Molecular mechanisms implicated in Thymic Stromal Lymphopoïetin-induced dendritic cell
migration
Carolina Martinez-Cingolani, Antonio Cappuccio, Vassili Soumelis
Inserm U932, Institut Curie, Paris, France.
Once activated, Dendritic Cells (DCs) migrate to the lymphoid organs and exert their role as antigen
presenting cells. Thymic stromal lymphopoietin (TSLP) is a pro-inflammatory cytokine, secreted by
inflamed skin and epithelia, which strongly activates myeloid DCs. The TSLP-activated DCs secrete
the inflammatory chemokines CCL17 and CCL22, prime an inflammatory Th2 response, and have
been involved in the pathogenesis of allergic inflammation. We recently showed that TSLP also
induces cytoskeleton polarization and myeloid DC migration. The aim of the present study is to
understand the mechanisms by which TSLP induces migration of human blood myeloid DC subsets.
To evaluate if TSLP induces DC migration in a direct way or through the secretion of chemokines
acting in an autocrine manner, we performed migration experiments using a specific anti-TSLP
neutralizing antibody and pertussis toxin (PTX), an inhibitor of GPCR signaling. We found that TSLPinduced secreted factors could not recapitulate DC migration. In addition, we found that the use of
PTX inhibited TSLP-induced migration. Therefore, TSLP is required to induce DC migration, but this
effect is dependent on the expression of a PTX-sensitive chemokine receptor. Moreover, we found
that TSLP-induced migration was restricted to the blood BDCA-1+ subset since BDCA-3+ subset did
not migrate in response to TSLP.
Our results highlight intricated mechanisms underlying TSLP-induced DC migration. Ongoing
transcriptomics analysis of TSLP-treated BDCA-1+ and BDCA-3+ DCs should shed light on the
molecular mechanisms involved and the TSLP-induced signaling pathways on blood DC subsets.
Notes:
44 12.15 – 12.30
October 12, 2013
SELECTED ABSTRACT
Immune complex-mediated recruitment of slanDCs and NK cells to vascular endothelium
Thomas Döbel, Anke Lonsdorf, Alexander Enk and Knut Schäkel
Heidelberg University Hospital, Department of Dermatology, Heidelberg, Germany
Several autoimmunity-driven inflammatory diseases with skin manifestations like lupus erythematosus
and vasculitis are associated with immune complex (IC) deposition in the vascular bed or antiendothelial cell antibodies. Fc receptor-based detection of such autoICs by immune cells leads to ICdependent inflammation and tissue damage. To investigate modes of immune cell recruitment to areas
of IC deposition we used a perfusion system coupled with time-lapse video microscopy and measured
arrest functions of various leukocytes on immobilized ICs. At flow conditions corresponding to surface
shear stress within human venous capillaries (>0.5 dynes/cm2) a pronounced recruitment of FcγRIII
(CD16) positive slanDCs and NK cells could be observed. On the other hand human plasmacytoid
DCs, CD1c+ DCs or Fc receptor negative control cells like T cells completely failed to adhere.
Investigating the receptor specificity of this IC-mediated attachment with blocking mAbs clearly
showed a dependence on CD16a while FcγRII (CD32) was largely dispensable. We performed
additional experiments with immobilized human IgG subtypes and recognized that especially IgG3
promoted a strong attachment of slanDCs, which was again largely dependent on CD16a. Finally,
when we performed experiments with monolayers of human dermal microvascular endothelial cells we
also found a pronounced CD16a-dependent recruitment of slanDCs to endothelial cells that were preincubated with endothelial cell-specific antibodies. Of note, this CD16a/IC-dependent recruitment at
the vascular interface is also supported by immunohistochemical identification of slanDCs in tissue
samples of lupus nephritis and allergic vasculitis. Collectively, our results show an efficient CD16amediated and shear stress-resistant adhesion of circulating slanDCs and NK cells on immobilized ICs.
These data provide evidence for a novel conduit of rapid FcR-dependent recruitment of immune cells
in IC-mediated tissue inflammation.
Notes:
October 12, 2013
12.30 – 12.45
Langerhans cells and Langerin+ dermal dendritic cells promote humoral responses through
the generation of follicular helper T cells
Chen Yao1, Elizabeth S. Jarrett1, Sandra M. Zurawski2, Gerard Zurawski2, Daniel H. Kaplan1, Botond
Z. Igyártó1
1
Department of Dermatology, Center for Immunology, University of Minnesota, Minneapolis, MN
55455, USA
2
Baylor Institute for Immunology Research and INSERM U899 - ANRS Center for Human Vaccines,
Dallas, TX 75204, USA
Langerhans cells (LC) are required for the induction of adaptive Th17 responses against Candida
albicans skin infection and for humoral immunity in an experimental staphylococcal scalded skin
model. In contrast, CD103+ Langerin+ dermal dendritic cells (dDC) are required for Th1 induction. To
study the effects of antigen presentation by LC in vivo under steady-state conditions, we immunized
transgenic mice that express human Langerin only in LC with anti-human Langerin/antigen complexes.
We also immunized LC-deficient mice with anti-murine Langerin/antigen complexes to selectively
target CD103+ Langerin+ dDC. In both cases DC remained immature and antigen was targeted
exclusively to either LC or CD103+ Langerin+ dDC. Steady-state presentation by either subset was
sufficient to promote proliferation of endogenous antigen-specific CD4 T cells as assayed by I-Ab
tetramer. The response peaked at day 7 without evidence of deletion. Both LC and CD103+ Langerin+
dDC induced a small, transient expansion of antigen-specific Foxp3+ cells that was absent by day 10.
In contrast, both subsets induced strong expansion of antigen-specific follicular helper T cell
phenotype (Tfh) that persisted into the memory phase. These cells expressed CXCR5, PD-1, Bcl6,
IFN-γ, IL-21 and IL-4. Antigen targeting was also associated with the formation of germinal centers in
regional LN and the development of a robust humoral response. Notably, Tfh expansion and the
magnitude of the humoral response was unchanged if mice were also infected with C. albicans. Thus,
both LC and CD103+ Langerin+ dDC efficiently promote Tfh and humoral responses irrespective of
their state of maturation. We speculate that this function may have evolved to promote neutralizing
immunity to recurrent viral skin infections.
Notes:
46 12.45 – 13.00
October 12, 2013
SELECTED ABSTRACT
Characterization of the intrinsic cross-presentation ability of human dendritic cell subsets
Elodie Segura, Melanie Durand, Sebastian Amigorena
Institut Curie, INSERM U932, 26 rue d’Ulm, 75005 Paris, France
Dendritic cells (DCs) represent a heterogenous population. In mice, lymphoid organ-resident CD8+
DCs are specialized at cross-presentation, due to specific adaptations of their endocytic pathways
(high pH, low degradation, high export to the cytosol). We investigated the intrinsic cross-presentation
ability of in vitro-derived dermal DC equivalents, CD1a+ and CD14+ DCs, and of ex vivo lymphoid
organ-resident BDCA1+ DCs, BDCA3+ DCs and pDCs. Only CD1a+ DCs, but not CD14+ DCs, crosspresent efficiently and display high pH in their phagosomes and a high capacity to transfer exogenous
proteins to their cytosol. By contrast, all resident DC subsets cross-present soluble antigen efficiently,
as compared to macrophages. In addition, BDCA1+ and BDCA3+ DCs display similar phagosomal pH
and similar production of reactive oxygen species in their phagosomes. All three resident DC subsets
also efficiently export internalized proteins to the cytosol. We conclude that, while intracellular crosspresentation mechanisms are conserved between mouse and human, the DC subset specialization for
cross-presentation differs between mouse and human.
Notes:
47 INVITED SPEAKER
October 12, 2013
Bertrand Dubois – INSERM, Lyon, France
Langerhans cells dictate skin reactivity to weak contact allergens
48 14.30 – 15.00
15.00 – 15.30
October 12, 2013
INVITED SPEAKER
Natalija Novak – University of Bonn Medical Center, Bonn, Germany
The role of human Langerhans cells of the oral mucosa and the skin in allergic
diseases
49 INVITED SPEAKER
October 12, 2013
Björn Clausen – University Medical Center, Mainz, Germany
Dissecting the dendritic cell network in psoriasis-like skin disease
50 15.30 – 16.00
16.30 – 17.00
October 12, 2013
INVITED SPEAKER
Matthew Collin – Newcastle University, Newcastle upon Tyne, UK
Langerin expression and mutated BRAF in human dendritic cells: more clues about
the origin of Langerhans cell histiocytosis
October 12, 2013
17.00 – 17.15
The role of dendritic cells in psoriasis
Elisabeth Glitzner1, Ana Korosec1, Martin Holcmann1, Barbara Drobits1, Helia Berrit Schönthaler2,
Erwin F. Wagner2, Maria Sibilia1
1
2
Institute of Cancer Research, Medical University of Vienna, 1090 Vienna, Austria
Centro Nacional de Investigaciones Oncológicas, 28029 Madrid, Spain
Psoriasis is a frequent, chronic disease in which the epidermis is affected by aberrant proliferation and
differentiation, accompanied by severe skin inflammation. Within psoriatic lesions the number of
Langerhans cells (LCs), the resident dendritic cells (DCs) of the epidermis, is often reduced compared
to non-psoriatic skin, whereas plasmacytoid DC (pDC) numbers are increased. The exact role of these
DC subtypes in psoriasis is, however, still poorly investigated. In this study we analyzed the
involvement of LCs and pDCs in disease initiation and progression by employing mice harboring an
inducible deletion of the AP-1 transcription factors c-jun and junB in the basal layer of the epidermis
(jun/junBΔep) that develop a psoriasis-like phenotype resembling human disease. LCs gradually
disappeared from the epidermis when psoriatic disease progressed, concomitant to their maturation
and migration to local draining lymph nodes. pDCs, that are normally absent from healthy skin,
immigrated into the dermis of jun/junBΔep mice in high numbers parallel to disease progression. To
study the function of these two DC subsets in psoriasis, we crossed jun/junBΔep mice to two transgenic
mouse strains where Langerin+ cells or pDCs could be inducibly depleted based on targeted
expression of the diphtheria-toxin receptor. When Langerin+ cells were depleted during active
disease, we found that disease symptoms were severely aggravated. In contrast, psoriatic disease
developed and progressed normally when Langerin+ cells were depleted before disease onset. Using
bone marrow chimeras between jun/junBΔep and Langerin-DTR mice we further dissected the
involvement of LCs and Langerin+ dermal DCs in psoriasis progression. Furthermore, depletion of
pDCs prior to disease induction resulted in a milder course of disease, whereas depletion of pDCs
during established disease had no impact. In summary, these results suggest that LCs have a
preventive role during active psoriatic disease, while pDCs exert an important instigatory function
during its initiation phase.
Notes:
52 17.15 – 17.30
October 12, 2013
SELECTED ABSTRACT
Interleukin 20 protein locates to distinct mononuclear cells in psoriatic skin
Rikke Bech1,2, Kristian Otkjaer1, Svend Birkelund3, Thomas Vorup-Jensen2, Ralf Agger3, Lars Iversen1,
Knud Kragballe1, John Rømer4
1
Department of Dermatology, Aarhus University Hospital, Aarhus, Denmark
Department of Biomedicine, Aarhus University, Aarhus, Denmark
3
Department of Health Science and Technology, Aalborg University, Aalborg, Denmark
4
Department of Histology, Novo Nordisk, Maaloev, Copenhagen, Denmark
2
We have previously demonstrated that mRNA for the pro-inflammatory cytokine interleukin 20 (IL-20)
is expressed in suprapapillary keratinocytes of lesional psoriatic skin. Here, we describe the
distribution of IL-20 protein and the identity of the IL-20-positive cells in lesional psoriatic skin. By
immunohistochemistry we found that the main part of IL-20 immunoreactivity is present in
mononuclear cells of the dermal papillae. Double-immunofluorescence studies revealed that the IL-20
positive cells located in the papillae were positive for Langerin, CD1a, CD4, and CD303, whereas they
were negative for CD11c and CD123. In order to clarify whether the IL-20 protein was actively
produced by or accumulated in these cells through uptake, we did in situ hybridization for IL-20 mRNA
on non-lesional psoriatic skin, cultured ex vivo and stimulated with IL-1β. We observed a colocalization between IL-20 mRNA and the keratinocyte marker CK14. Importantly, no IL-20 mRNA was
detected in the dermal mononuclear cells. Our results suggest that IL-20 is produced by keratinocytes,
released into the epidermis, and then possibly taken up by papillary mononuclear cells. Based on the
co-localisation studies we suggest that the IL-20 positive mononuclear cells in the papillae are
immature dendritic cells. Neutralization of IL-20 may be beneficial in patients with psoriasis.
Notes:
October 12, 2013
17.30 – 17.45
Transcriptional profiling of human Langerhans cells in tumor draining sentinel lymph nodes
Rieneke van de Ven1,2, Christopher Dubay1, Malin Lindstedt3, Bernard A. Fox1 and Tanja D. de Gruijl2
1
Laboratory of Molecular and Tumor immunology, Earle A. Chiles Research Institute, Portland,
Oregon, USA
2
Department of Medical Oncology, VU University medical Center-Cancer Center Amsterdam,
3
Department of Immunotechnology, Lund University, Lund 22363, Sweden
Langerhans cells (LC) and dermal dendritic cells (DDC) that migrate from the skin to the draining
lymph nodes upon encounter of antigens are likely key orchestrators of an induced immune response
against these antigens. We hypothesize that when a tumor shares draining lymph nodes (LN) with
proximate skin, as is the case for melanoma or breast tumors, the tumor microenvironment will impact
the gene expression profile in these skin-derived DC. For this study, we isolated LC as well as CD1a+
DDC and two non-skin derived LN-resident DC subsets from metastasis-negative sentinel lymph
nodes (SLN) from two breast cancer patients. After RNA isolation, the samples were hybridized on
Affymetrix human U133 Plus 2.0 arrays. We were able to compare the gene expression profile of
these LC and DDC with LC and DDC isolated from healthy donor human skin, on which we had
previously conducted a similar gene array analysis. Initial analysis shows that many pathways that are
highly over-expressed in SLN-LC compared to healthy skin-LC are tumor-related pathways and
include elevated expression of immune inhibitory molecules such as IDO-1/2, Neuropilin-2, STAT3,
PD-L1, PD-L2 and TGF-beta2, while maturation markers such as CD86, CD83 and HLA-DR were
reduced. As a biological control, the data show that Langerin and CD1a expression are significantly
reduced on SLN-LC compared to skin-LC, a process known to happen upon migration from the skin to
the nodes. Interestingly, the cytokine IL-15, which is known for its importance for CD8 T-cell priming by
LC, is increased in the LC present in the lymph nodes, the site where they would most likely encounter
T cells. This unique set of data will provide valuable clues to further unravel functional differences
between different primary human DC subsets in tumor-draining SLN as well as between pre- and postmigration LC and DDC.
Notes:
54 17.45 – 18.00
October 12, 2013
SELECTED ABSTRACT
Langerhans Cells suppress innate skin inflammation via liver CXCR6+ NK cells
Felix Scholz, Daniel H. Kaplan
Department of Dermatology, Center for Immunology, University of Minnesota, Minneapolis, MN 55455,
USA
The skin serves as a barrier to protect the organism from physical damage, dehydration and infection.
In addition to a physical barrier, the skin-resident cells recognize pathogens and rapidly recruit innate
immune effectors. We found that intradermal footpad injection of a fungus, C. albicans, or a gram
positive bacteria resulted in a robust inflammation associated with an influx into the foot pad of
monocytes and neutrophils within 4 hours in naïve mice that resolved within 48 hours. This
inflammation was reduced in MyD88-/- and IL-6-/- mice but was unaffected in Rag1-/- mice. Notably,
we observed that footpad inflammation and levels of IL-6 in huLangerin-DTA mice that lack
Langerhans cells (LC) were exaggerated compared to litter mate controls. The exaggerated
inflammation was independent of adaptive immunity since we observed a similar degree of
inflammation in huLangerin-DTA x Rag1-/- mice. In contrast, antibody-mediated ablation of natural
killer (NK) cells with either anti asialo-GM1 or anti-NK1.1 reversed the phenotype. Recently CXCR6+
NK cells have been described as a liver-resident subset that mediate antigen-specific “memory like”
responses during contact hypersensitivity. We found that exaggerated inflammation was ameliorated
in huLangerin-DTA x CXCR6-/- mice. Moreover, adoptive transfer of liver lymphocytes from
huLangerin-DTA but not control mice into wild type mice reproduced the exaggerated inflammation.
Thus, in the absence of LC, NK1.1+ CXCR6+ NK cells become dysregulated and promote
exaggerated innate immune responses to fungal and bacterial pathogens in the skin. This suggests
novel model in which LC directly or indirectly suppress inappropriate activation of “memory-like” NK
cells.
Notes:
October 12, 2013
18.00 – 18.15
Langerhans cell homeostasis and activation is altered in epidermis expressing human
papillomavirus type 16 E7
Nor Malia Abd Warif1, Patrizia Stoitzner2, Graham R. Leggatt1, Stephen R. Mattarollo1, Ian H. Frazer1
and Merilyn Hibma3
1
University of Queensland Diamantina Institute, Princess Alexandra Hospital, Ipswich Road,
Woolloongabba, Brisbane, Qld 4102, Australia
2
Department of Dermatology and Venereology, Medical University Innsbruck, Innsbruck, Austria
3
Department of Microbiology and Immunology, University of Otago, Dunedin, New Zealand
The incidence of squamous cell carcinoma and its pre-malignancies is increasing, however little is
known of Langerhans cell (LC) regulation in premalignant skin. Human papillomavirus (HPV) type 16
E7 is a cell cycle deregulating protein that contributes to the oncogenesis of HPV16. E7-expressing
skin from the K14E7 mouse is hyperplastic, hyperkeratotic and contains inflammatory infiltrates.
Despite the different etiology, these characteristics parallel squamous cell pre-malignancy. The
purpose of this study is to determine if the frequency and function of LC is altered in hyperplastic
K14E7 mouse skin.
LC number and function was compared between the K14E7 mouse and control C57Bl/6 mice. We
show that number of resident LC in the skin of the K14E7 was more than doubled. In contrast,
migration of LC from the K14E7 skin was approximately halved. Functionally, a greater percentage of
LC from the K14E7 epidermis was competent to take up antigen and those cells took up more antigen
than LC from control epidermis. In addition, MHC class I and II and the costimulatory molecule
expression was increased on LC from the K14E7 epidermis. Of those markers, only expression of
MHC class II was dependent on the presence of functional T cells. When extracted from the skin and
matured, LC from the K14E7 mouse did not differ from control LC in their ability to present or cross
present antigen to T cells.
The increased number, activation status and enhanced antigen uptake of the LC is consistent with
their being regulated in the microenvironment of the hyperplastic K14E7 skin. This coupled with their
reduced migration suggests that the LC retained in the 14E7 skin may function to regulate the local
skin-resident T cells.
Notes:
56 18.15 – 18.30
October 12, 2013
SELECTED ABSTRACT
Technical advance: anti-CXCL12 inhibits mature LC migration in immuno-competent skin
equivalents exposed to allergens
Ilona J. Kosten, Sander W. Spiekstra, Tanja D. de Gruijl, Sue Gibbs
Department of Dermatology VUmc Amsterdam, The Netherlands
Department of Medical Oncology VUmc Amsterdam, The Netherlands
Academic Centre for Dentistry (ACTA) Amsterdam, The Netherlands
Here we describe the first immuno-competent full thickness human skin equivalent model
which can be used to investigate both mechanisms involved in LC maturation and migration
after exposure to allergens or to irritants.
We have made use of a novel tissue-engineered immuno-competent skin equivalent (SE)
model that structurally and functionally resembles native human skin. It consists of a
reconstructed epidermis containing human keratinocytes and Langerhans-like cells derived
from the human MUTZ-3 cell line (MUTZ-LC) on a human fibroblast-populated dermis. The
CD1a+ MUTZ-LC populate the entire epidermis at a similar density to that found in native
skin. Exposure of the SE to sub-toxic concentrations of the allergens NiSO4, Cinnamaldehyde,
Resorcinol and the irritants SDS and TritonX resulted in LC migration out of the epidermis
towards the fibroblast populated dermal compartment.
A significant dose-dependent upregulation of the DC maturation-related genes CCR7 and
IL-1β upon epidermal exposure to the allergens was found, indicative of maturation and
migration of the epidermally incorporated LC. IL-1β was already upregulated in the
epidermis, whereas CCR7 was only upregulated in the dermis, indicating that CCR7
upregulation may occur under influence of dermal fibroblasts. Neutralizing antibody to
CXCL12 blocked the allergen induced migration, but not the irritant induced migration in a
similar manner to that previously shown using intact ex vivo human skin. This immunocompetent
human skin model provides a unique reproducible research tool to study human
Langerhans cell biology in situ under controlled in vitro conditions.
Notes:
57 INVITED SPEAKER
October 13, 2013
Béhazine Combadière – INSERM, Paris, France
Langerhans cell's pivotal role in tailored immunity to vaccines
58 09.00 – 09.30
09.30 – 10.00
October 13, 2013
INVITED SPEAKER
Yvette van Kooyk – Free University Medical Center, Amsterdam, The Netherlands
Glycan-based targeting of vaccines to Langerhans cells and dendritic cells in the skin
October 13, 2013
10.00 – 10.15
Crosstalk between human DC subsets promotes anti-mycobacterial activity and CD8 T cell
stimulation in response to tuberculosis vaccine Bacille Calmette–Guérin
Laura Lozza1, Maura Farinacci1, Kellen Fae1, Marina Bechtle1, Anca Dorhoi1, Mario Bauer2, Franca del
Nonno3, Stefan H.E. Kaufmann1
1
Max Planck Institute for Infection Biology, Department of Immunology, Berlin, Germany
Helmholtz Centre for Environmental Research – UFZ, Department of Environmental Immunology
3
Pathology Division, National Institute for Infectious Disease ‘L. Spallanzani’, Rome, Italy
2
Tuberculosis (TB) remains a global health threat caused by the intracellular bacterial pathogen
Mycobacterium tuberculosis (Mtb). The only currently available vaccine against TB is an attenuated M.
bovis, called bacille Calmette–Guérin (BCG). Vaccine efficacy against TB implies the activation of an
efficient immune response with dendritic cells (DCs) playing a central role. To date little is known
about how specialized human DC subsets contribute to protection against TB. We analyzed the role of
human pDCs, BDCA-1+ mDCs and BDCA-3+ mDCs in the immune response to BCG or Mtb. Since
pDCs can be activated by bystander stimulation the mutual interaction of pDCs and mDC subsets in
response to BCG was also analyzed. We found that pDCs are activated by BCG-infected BDCA-1+
mDCs to upregulate maturation markers and to produce granzyme B, but not IFN-α. Reciprocally, the
presence of activated pDCs enhanced IL-1β production and mycobacterial growth control by infected
mDCs. Synergy between the unique capacities of the two DC subsets promoted BCG-specific CD8 T
cell stimulation. BDCA-3+ mDCs were also susceptible to BCG infection but less than BDCA-1+ mDCs
and failed to prime CD8 T cells in our system. Contact with Mtb-infected mDCs induced GrB-pDC
activation indicating that pDCs responded to the state of activation of BDCA-1+ mDCs rather than to
bacterial species of different virulence. Consistent with these data, we identified discrete contact areas
between mDCs and pDCs and the presence of GrB-pDCs in lymph nodes of TB patients. We conclude
that mDC–pDC cross-talk should be exploited for rational design of BCG-based next-generation TB
vaccines.
Notes:
60 10.15 – 10.30
October 13, 2013
SELECTED ABSTRACT
Efficient cross-priming and Th1 induction by Langerhans cells following microneedle-mediated
nanoparticle transdermal vaccination
Marija Zaric1, Oksana Lyubomska1, Christopher J. Scott2, Ryan F. Donnelly2, Adrien Kissenpfennig1
1
The Centre for Infection and Immunity, School of Medicine, Dentistry and Biomedical Sciences,
Queen’s University Belfast, University Road, Belfast, BT9 7AE, UK.
2
School of Pharmacy, Queen’s University Belfast, Lisburn Road, Belfast, BT9 7BL, UK
Harnessing dendritic cells (DCs) for development of effective vaccines necessitates further
understanding of the functional specialization of distinct DC populations in vivo. We used polymeric
dissolvable microneedle (MN) arrays laden with antigen encapsulated nanoparticles (NPs) to target
skin DCs networks in mice and to investigate the capacity of different skin DCs subsets to present
antigen to antigen-specific naïve T cells. We demonstrate that although all skin DC subsets efficiently
take up and deliver NPs to the skin draining lymph nodes, Langerhans cells (LCs) constitute the major
subset capable of antigen cross-presentation to CD8+ T cells. Indeed, we show that LCs are critical
for effective immunization using antigen-laden NPs, as depletion of LCs dramatically reduces the
proliferation of antigen-specific CD8+ T cells in vivo. LCs are also necessary for IFN-γ and IL-17
production by antigen-specific CD4+ T cells. Moreover, we have explored the contribution of skin DC
subsets for the generation of antigen-specific anti-tumor and anti-viral immune responses following
MN immunization in vivo and demonstrate that depletion of LCs significantly reduces protective
immunity in both disease models. Therefore, targeting of nano-encapsulated antigen to specific skin
DC subsets, in particular to LCs, through dissolvable MNs potentially provides a promising
technological platform for improved vaccination efficacy through enhanced cytotoxic T lymphocyte
responses.
Notes:
61 INVITED SPEAKER
October 13, 2013
Adrien Kissenpfennig – Queens University Belfast, Belfast, Uk
Can Langerhans cells take the sting out of vaccines?
62 11.00 – 11.30
11.30 – 11.45
October 13, 2013
SELECTED ABSTRACT
Targeting of myelin oligodendrocyte glycoprotein to the DC/LC antigen receptor DEC205 in
vivo induces immune suppression and prevents allergic experimental encephalomyelitis (EAE)
in mice
Sabine Ring, Michael Maas, Alexander Enk, Karsten Mahnke
Ruprecht-Karls-University Heidelberg, Department of Dermatology, Vosstrasse 11, D-69115
Heidelberg, Germany
The antigen receptor DEC-205 is expressed by dendritic cells (DC) and greatly increases antigen
presentation over pinocytosis or phagocytosis. After injection of antigens coupled to antibodies specific
for DEC-205 into mice, effective presentation of the antigens to T cell follows. When antigen
presentation occurs by non-activated “steady state” DC in vivo, we observed induction of regulatory T
cells. We created single chain fragment variables (scFv) specific for DEC-205 fused to the self antigen
MOG, to target non-activated DC in vivo and to induce tolerance to EAE.
DEC-scFv:MOG fusions proteins, as well as isotype controls were expressed and purified from E.coli
and immunohistochemical staining of CD11c+ dendritic cells displayed a positive staining for scFv,
colocalizing with MHC class II. For functional testing DEC-scFv:MOG was injected into mice and DC
were analyzed 3 days later. We found that DC isolated from DEC-scFv:MOG injected mice stimulated
vigorous proliferation of MOG-specific 2D2 T cells, indicating presentation of the relevant MOG
peptide in vitro. Furthermore, DC isolated from those animals produced significantly more TGF-β as
well as IL-10, as compared to isotype-treated or untreated mice, respectively. And when analyzing the
T cell compartment we recorded elevated numbers of activated CD4+CD25+FoxP3+ Treg (16% of
CD4) in the spleen after injection of DEC-scFv:MOG as compared to controls (12% of CD4). These
Treg produced significantly more IL-10 as compared to controls. Most importantly, when EAE was
induced in DEC-scFv:MOG-injected mice and in isotype-scFv:MOG treated controls, none of the DECscFv:MOG injected mice displayed any EAE symptoms in contrast to controls, which developed
severe EAE. In a therapeutical setting, EAE was induced first and after mice had developed mild
symptoms, they were treated with respective scFv conjugates. Here injection of DEC-scFv:MOG lead
to abrogation of the disease in >90% of the animals tested. In contrast, all animals in the control
groups developed a severe EAE.
These data indicate that targeting of MOG to steady state DC in vivo prevents EAE by a DC/Tregdriven mechanism.
Notes:
October 13, 2013
11.45 – 12.00
C-type lectin mediated uptake of glycan modified antigens by primary human Langerhans cells
results in cross presentation to CD8+ T cells
Cynthia M. Fehres1, Sven C.M. Bruijns1, Martino Ambrosini1, Hakan Kalay1, Sandra J. van Vliet1, Erik
Hooijberg2, Tanja D. de Gruijl3, Wendy W.J. Unger1 and Yvette van Kooyk1
1
Department of Molecular Cell Biology and Immunology, VU University Medical Center
Department of Pathology, VU University Medical Center
3
Department of Medical Oncology, VU University Medical Center
2
The potential of the skin immune system to generate humoral and cellular immune responses is well
established and the skin is actively exploited as vaccination site. However, the precise function of the
skin dendritic cell (DC) subsets, such as epidermal Langerhans cells (LCs) and dermal CD1a+ and
CD14+ DCs, within the skin immune system is controversial and still under debate. Especially the
capacity of each subset to cross-present exogenous delivered antigens is unclear, but of particular
importance in the design of effective immunotherapies. Here, we show that primary human LCs crosspresent MART-1 peptide more efficiently than the dermal DCs. Modification of MART-1 using
antibodies to the C-type lectins Langerin and DEC-205, but not Dectin-1, further enhanced the crosspresenting capacity of the LCs. Additionally, coupling of glycans, which are recognized by Langerin, to
MART-1 peptides resulted in Langerin-mediated uptake and enhanced cross-presentation, indicating
that antigens taken up via Langerin route to HLA class I loading compartments. The potency to
enhance CD8+ T cell responses could be further increased through activation of LCs with the TLR3
ligand polyI:C. Our results provide a rationale for the development of LC-targeting immune therapies,
through glycans or antibodies, in order to generate effective anti-tumor or anti-viral CTL responses.
Notes:
64 12.00 – 12.30
October 13, 2013
INVITED SPEAKER
Niki Romani – Innsbruck Medical University, Innsbruck, Austria
Wrapping up LC2013: the current status of Langerhans cells, their future and beyond
65 66 POSTER PRESENTATIONS
67 POSTER PRESENTATIONS
P1.
Distinctive antibody responses induced by immunization of mice with rat monoclonal
antibodies specific for a panel of dendritic cell surface markers
L.H. Pugholm, L.R. Petersen, K. Varming and R. Agger
P2.
MUTZ-3-derived Langerhans-like cells as a model system to study cytokine- and TLR2induced dendritic cell activation
Stephanie Bock, André Said, Günther Weindl
P3.
Human epidermal Langerhans cells co-express indoleamine 2,3-dioxygenase and CD83
Paola Di Gennaro, Maria Raffaella Romoli, Gianni Gerlini, Massimo D’Amico, Paola Brandani,
Nicola Pimpinelli, Lorenzo Borgognoni
P4.
Recognition of fungal pathogens instructs human Langerhans cells to induce Th1
responses
Agata Drewniak, Marein A.V.P de Jong, Sonja I. Gringhuis, Teunis B. Geijtenbeek
P5.
The psoriasis-associated cytokine interleukin-23 inhibits Langerhans’ cell migration
Laura H Eaton, Rebecca J Dearman, Christopher EM Griffiths, Ian Kimber
P6.
Skin Langerhans cell histiocytosis in a child. A case report
Rossella Filippetti and Claudia Canofori
P7.
Integrated study of dermal CD11b+ mononuclear phagocytes including conventional
and monocyte-derived dendritic cells and macrophages present in the mouse skin
Samira Tamoutounour, Martin Guilliams, Frederic Montanana Sanchis, Dora Terhorst, Camille
Malosse, Marc Dalod, Bernard Malissen and Sandrine Henri
P8.
Could dermal dendritic cells play a role in the development of neutralizing anti-drug
antibodies?
Christina Hermanrud, Anna Fogdell-Hahn
P9.
Casein kinase II regulates the recycling of the DC/LC antigen receptor DEC205 from
MHC class II+ compartments back to the cell surface
Rainer Koch, Sabine Ring, Alexander H.Enk, Karsten Mahnke
P10.
Characterization of anti-tumoral immune responses in a mouse model of spontaneous
melanoma
David G Mairhofer, Vincent Flacher, Suzie Chen, Juergen C Becker and Patrizia Stoitzner
P11.
EFdA, a reverse transcriptase inhibitor, potently blocks HIV-1 ex vivo infection of
Langerhans cells within epithelium
Takamitsu Matsuzawa, Tatsuyoshi Kawamura, Youichi Ogawa, Kohji Moriishi, Yoshio
Koyanagi, Hiroyuki Gatanaga, Shinji Shimada, and Hiroaki Mitsuya
P12.
Differential cytokine requirements for oxazolone and DNCB for allergen-induced
Langerhans’ cell migration
Aleksandra Metryka, Laura H Eaton, Ian Kimber, Ruth A Roberts and Rebecca J Dearman
P13.
Revisiting the origin of Langerhans cell histiocytosis: mutated BRAF gene identifies
potential circulating precursor cells
Paul Milne, Venetia Bigley, Naomi McGovern, Matthew Collin
P14.
The role of langerin-positive skin dendritic cells in skin carcinogenesis
Daniela Ortner, Christoph Tripp, Nicole Amberg, Maria Sibilia, Björn E. Clausen and Patrizia
Stoitzner
P15.
HIV infections in novel CD1a cells from human vaginal mucosa and epidermal
Langerhans cells
Victor Peña-Cruz, Rahm Gummurulu and Manish Sagar
68 POSTER PRESENTATIONS
P16.
Distinct molecular signature of human skin Langerhans cells denotes critical
differences in cutaneous dendritic cell immune regulation
Marta E Polak, Stephen M Thirdborough, Chuin Y. Ung, Tim Elliott, Eugene Healy, Tom C
Freeman, Michael R Ardern-Jones
P17.
Poly(I:C) induces Th1 cytokines and IFNbeta in monocyte-derived Langerhans-like cells
through TLR3
André Said, Monika Schäfer-Korting, Günther Weindl
P18.
Human Langerhans cells play a crucial role in CCR5 tropic selection of HIV-1 variants
during sexual transmission
Ramin Sarrami-Forooshani, Annelies W. Mesman, Carla M.S. Ribeiro, Michiel van der Vlist
and Teunis B. H. Geijtenbeek
P19.
Targeting the glutamate pathway: a potential approach for immunotherapy in
melanoma
Sandra C. Schaffenrath, Daniela Ortner, Suzie Chen, Nikolaus Romani and Patrizia Stoitzner
P20.
Novel hollow microneedle technology for depth controlled microinjection-mediated
dermal vaccination: a study with polio vaccine in rats
Koen van der Maaden, Bas Trietsch, Pim Schipper, Juha Monkare, Heleen Kraan, Eleni Maria
Varypataki, Stefan Romeijn, Raphäel Zwier, Heiko van der Linden, Gideon Kersten, Thomas
Thomas Hankemeier, Wim Jiskoot, Joke Bouwstra
P21.
CD14+ DDCs take the stage in allergen-specific intradermal immunotherapy
Na Luo, Yuri Souwer, Fong Lin, Ghaith Bakdash,Toni M van Capel, Martien L Kapsenberg,
Marcel BM Teunissen and Esther C de Jong
P22.
Dengue receptor CLEC5A induces immune responses in skin Langerhans cells
Joris K. Sprokholt, Sonja I. Gringhuis, Teunis B.H. Geijtenbeek
P23.
Polycomb Group Protein Ezh2 Regulates Integrin-Dependent Leukocyte Migration and
Methylation of Talin
M. Gunawan, N. Venkatesan, W.H. Neo, J. Li, J.T. Loh, J.F. Wong, T. Guo, C.E. See, S.
Yamazaki, K.C. Chin, L.G. Ng, K.Z. Sze, F. Ginhoux, I-H. Su
P24.
Unique role of Cbfb2 variant in Langerhans cell development
Mari Tenno, Yoshinori Naoe, Sawako Muroi and Ichiro Taniuchi
P25.
Characterization of dendritic cell subtypes in human melanoma and non-melanoma
skin cancer
Christoph H. Tripp, Van Anh Nguyen, Dietmar Heiser, Barbara Del Frari, Nikolaus Romani,
Patrizia Stoitzner
P26.
Activated vaginal Langerhans cells efficiently transmit HIV-1 to T cells
Nienke H. van Teijlingen, Carla M.S. Ribeiro, Elisabeth van Leeuwen, Joris A.M. van der Post,
Teunis B.H. Geijtenbeek
P27.
Innate dendritic cell sensing of opsonized HIV-1 to activate Th17 cells
Wilfried Posch, Andrea Schroll, Hubert Hackl, Cornelia Lass-Flörl, Gianfranco Pancino, Asier
Saez-Cirion, Zlatko Trajanoski, Teunis Geijtenbeek, Günter Weiss, Doris Wilflingseder
P28.
Protease inhibitor from tick saliva acts on human lysosomal enzymes in differentiated
MUTZ-3 cells
Tina Zavašnik-Bergant, Lenka Grunclová, Jiří Salát, Robert Vidmar, Petr Kopáček, Boris Turk
69 POSTER PRESENTATION
P1.
Distinctive antibody responses induced by immunization of mice with rat monoclonal
antibodies specific for a panel of dendritic cell surface markers
L.H. Pugholm1,2, L.R. Petersen2, K. Varming1 and R. Agger2
1
Department of Clinical Immunology, Aalborg University Hospital, Aalborg, Denmark
Laboratory of Immunology, Department of Health Science and Technology, Aalborg University,
Aalborg, Denmark
2
During recent years experimental evidence has been published demonstrating that targeting antigen
(Ag) to surface markers on dendritic cells (DCs) may be a very efficient way of inducing immune
responses. These results have aroused hopes that it may be possible to develop DC-targeting
vaccines that effectively prevent or treat human diseases by inducing immune responses that are both
potent and suitably tailored to the given situation. A number of DC surface molecules have been
investigated for their usefulness as targets for antigen-delivery, but many more deserve examination.
In the present study, we compared antibody responses in the mouse following targeting of antigen to a
series of DC surface markers. Taking advantage of the fact that rat monoclonal antibodies (mAbs)
against murine DCs may act as both targeting devices (the Ag-binding parts of the molecule) and Ag
(epitopes on the rat immunoglobulin that are immunogenic in the mouse), groups of mice were
vaccinated subcutaneously, in the absence of adjuvant, with mAbs against the following markers:
CD11c, CD36, CD205, CD206, CD209, Clec6A, Clec7A, Clec9A, Siglec H and PDC-TREM. Control
groups were vaccinated with non-targeting, isotype-matched mAbs. The serum levels of antibodies
against rat IgG, and the IgG subclass composition of the responses, were monitored by ELISA for 84
days following vaccination. Experiments elucidating the influence of the route of vaccination were also
performed.
From the results it is evident that several different surface molecules on murine DCs may function as
efficient targets for the induction of antibody responses. Targets that elicited strong antibody
responses were PDC-TREM, CD11c, CD36, CD205, Clec6A and Clec7A, while targeting CD206,
CD209, Clec9A and Siglec H, resulted in poor responses, comparable to those elicited by nontargeted mAbs. Data on the effect of the route of administration will be presented on the poster.
Notes:
P2.
MUTZ-3-derived Langerhans-like cells as a model system to study cytokine- and TLR2induced dendritic cell activation
Stephanie Bock, André Said, Günther Weindl
Langerhans cells (LC) represent a highly specialized subset of epidermal dendritic cells (DC), yet not
fully understood in their role of balancing skin immunity. To study human epidermal DC behavior, we
used in vitro-generated Langerhans-like cells derived from peripheral blood monocytes (MoLC) as well
as from the acute myeloid leukemia cell line MUTZ-3 (MUTZ-LC). Comparative characterization of
MoLC and MUTZ-LC included phenotypic properties and constitutive toll-like receptor (TLR) and
sphingosine-1-phosphate receptor (S1PR) expression. Compared to MoLC, MUTZ-LC overall display
a less immature state and lack typical features of human epidermal DC (CD324, TROP-2, Axl) but
express high levels of CD1a, CD207 and CCR6 and hence show migration capacity induced by
CCL20. MUTZ-LC express high mRNA levels of TLR1, 2 and 6 and S1PR2 and 4. In contrast, TLR1, 3
and 8 and S1PR1 and 2 are the most highly expressed genes in MoLC. The functional properties of
MUTZ-LC were investigated after stimulation with different TLR ligands, pro-inflammatory cytokines
and soluble CD40 ligand (CD40L). Consistent with the TLR expression pattern, MUTZ-LC respond
robustly to TLR2/1 and TLR2/6 engagement with increased CD83 and CD86 protein expression as
well as upregulated IL-6, IL-12p40, IL-23p19, CCR7 and decreased CCR6 mRNA levels. Similar
results were obtained upon stimulation with pro-inflammatory cytokines whereas ligands to TLR3,
TLR4 and CD40L failed to activate MUTZ-LC. Moreover, CCR7-mediated migration towards CCL21
for MUTZ-LC correlated with their maturation status but revealed lower migratory capacity when
compared to MoLC. Taken together, our results support and extent previous findings that MUTZ-LC
represent a valuable model to study DC migration. MUTZ-LC furthermore may be particularly suitable
to study TLR2/1/6-mediated signaling involved in the regulation of LC function.
Notes:
P3.
Human epidermal Langerhans cells co-express indoleamine 2,3-dioxygenase and CD83
Paola Di Gennaro1,2*, Maria Raffaella Romoli2*, Gianni Gerlini1, Massimo D’Amico3, Paola Brandani1,
Nicola Pimpinelli2, Lorenzo Borgognoni1
1
Unit of Plastic and Reconstructive Surgery - Regional Melanoma Referral Center, Tuscan Tumour
Institute (ITT), Santa Maria Annunziata Hospital, Florence, Italy
2
Section of Dermatology, Dept. Translational Surgery and Medicine, University of Florence, Italy
3
Dept. Clinical and Experimental Medicine, University of Florence, Florence, Italy
* Authors contributed equally
Langerhans cells (LCs), the dendritic cells (DCs) of epidermis and mucosae, are characterized by
Langerin expression, a C-type lectin involved in antigen up-take and Birbeck granule formation. LCs
within epidermis are the typical immature DCs: weakly positive for HLA-DR and the co-stimulatory
molecule CD86 but negative for CD80 and the maturation marker CD83. Following activation by proinflammatory cytokines, LCs migrate to regional LNs reaching a full maturation state characterized by
HLA-DR and CD86 up-regulation, and CD80 and CD83 expression. Because of their localization, LCs
are the first DCs interacting with skin cancers, thereby they are thought to have a key role in tumour
immunology. For long time it has been thought that mature LCs activate immunity whereas immature
LCs induce tolerance. However it has been reported that freshly-isolated epidermal LCs, stimulated
with IFN-γ, expressed the tolerogenic enzyme indoleamine 2,3-dioxygenase (IDO), the regulator of
peripheral and tumour-induced tolerance. Thus the relation between IDO expression and LCs
maturation is a hot topic for cancer immunotherapy.
To address this issue we analyzed IDO and CD83 co-expression in immature and mature epidermal
LCs, using Langerin as track molecule and a well-established assay to induce maturation without the
addition of pro-inflammatory cytokines, in order to mimic what happens in vivo by LC migration to
lymph nodes upon activation.
By immunofluorescence analyses, in situ epidermal LCs were CD83- and did not express IDO, but
IDO+ cells were observed within migrated CD83+ LCs from epidermal explants. Quantitative threecolour flow cytometry analyses confirmed that freshly-isolated LCs were CD83- and IDO-, while only a
fraction of migrated CD83+ LCs expressed IDO (16.87 ± 7.63%, range 10.06 - 25.11%).
This is the first quantitative analysis of IDO and CD83 co-expression in epidermal Langerin+ LCs.
Given the tolerogenic role of IDO, these findings might indicate a possible regulatory role of IDO+
mature LCs.
Notes:
P4.
Recognition of fungal pathogens instructs human Langerhans cells to induce Th1
responses
Agata Drewniak, Marein A.V.P de Jong, Sonja I. Gringhuis, Teunis B. Geijtenbeek
Department of Experimental Immunology, Academic Medical Center, Amsterdam, The Netherlands
Fungal spices are ubiquitous residents of human skin and may cause invasive infections especially in
immune compromised individuals. Langerhans cells (LCs) are the subset of dendritic cells (DCs) that
line the epidermis, the outer layer of the human skin, and mucosal tissues. They function as antigen
presenting cells able to sense foreign antigens in the environment. Upon capture of antigen they are
able to migrate into the lymph node to instruct T cells to elicit appropriate immune response.
Here we analyze the ability of human skin-derived Langerhans cells to respond to fungal infection.
Human epidermal LCs are equipped with various receptors important for fungal recognition e.g. dectin1, TLR-2, langerin. Migratory LCs were able to bind and engulf various fungi, including C.albicans in a
langerin-dependent manner. Unstimulated LCs induce a potent Th2 response. Activation of LCs with
different strains of C.albicans led to induction of Th1 response, which was not observed in case of
stimulation with single-receptor ligands, such as curdlan. This Th1 skewing capacity was solely
dependent on IL12 production, as addition of neutralizing antibodies reversed this effect. These data
suggest that LCs are strong mediators of Th2 responses but upon fungal infection they are able to
induce Th1 responses required for antifungal immunity.
Notes:
P5.
The psoriasis-associated cytokine interleukin-23 inhibits Langerhans’ cell migration
Laura H Eaton1, Rebecca J Dearman1, Christopher EM Griffiths2, Ian Kimber1
1
The Faculty of Life Sciences, University of Manchester, Manchester, UK
The Dermatology Centre, University of Manchester, Manchester Academic Health Science Centre,
Manchester, UK
2
Psoriasis is an immune-mediated inflammatory disease affecting approximately 2% of the population.
We have shown previously that Langerhans’ cell (LC) migration from the epidermis is impaired in
patients with early-onset (presenting before age 40y) chronic plaque psoriasis. Psoriasis is associated
with increased expression of interleukin (IL)-23 and antibody therapy directed to the cytokine is an
effective therapy. Repeated intradermal injection of IL-23 in mice results in a skin phenotype
resembling psoriasis. The aim of these experiments was to investigate the role of IL-23 in the
regulation of LC mobilisation.
BALB/c strain mice received intradermal injections of 50 ng of IL-23 (or bovine serum albumin [BSA]
vehicle control) to the dorsum of each ear. In some experiments, 2 hours later, 25μl of 0.5% oxazolone
(a chemical contact allergen known to induce LC migration) was applied to the same site. Naïve mice
remained untreated. Four hours after oxazolone application (or 4 hours after IL-23 injection in groups
that did not receive oxazolone), mice were terminated, ears removed and the epidermis was isolated
and stained for LC. The frequency of LC in epidermal sheets was determined by fluorescence
microscopy. Percentage migration was calculated as the mean percentage loss of LC in the treated
compared with the concurrent control group.
Treatment with IL-23 alone did not induce significant migration; indeed there was evidence of
increased LC frequency from baseline (852 ± 18 to 957 ± 23 LC/mm2, p<0.01). Consistent with
previous observations, oxazolone exposure (after prior injection with BSA) resulted in migration of the
proportion of LC that appear to be available for mobilisation (15%; p<0.01). However, prior treatment
with IL-23 inhibited oxazolone-induced migration and LC frequencies were equivalent to those in naive
controls (942 ± 29 and 918.2 ± 24 LC/mm2, respectively). Thus, IL-23 inhibits LC mobilisation which
provides a potential mechanism for the observed impairment of LC migration in psoriasis skin.
Notes:
P6.
Skin Langerhans cell histiocytosis in a child. A case report
Rossella Filippetti and Claudia Canofori
Ospedale San Camillo Forlanini, Rome, Italy
Langerhans cell(LC) histiocytosis is a rare disease characterized by monoclonal proliferative disorder
of cells derived from bone marrow.
LC are located in epithelial surfaces, such as the skin, where their primary function, after they migrate
to regional lymph nodes, is to present foreign antigens to T cells, thus helping to initiate the adaptive
immune response.
The disease may be manifested as multiorgan-system involvement (often a relentless progressive
process with a poor prognosis)or as single- organ-system with a high likelihood of complete
spontaneous resolution.
Here we report a case of a 12 year old boy evaluated in our Day Service of Dermatology.
The patient presented with a 2 week history of cutaneous erosions and papule each 3 to 7 mm in
diameter with a hard serosanguineous crust as well as erythema scattered over lower trunk and
leg.The patient was afebrile,and the results of physical examination were normal.as the routine
laboratory tests.(viral,fungal cultures negative).
In the diagnostic work up a LC disorder was considered.
Skin biopsy is required for a definitive diagnosis with care taken to include both normal and lesional
skin in the specimen.
Microscopical examination revealed epidermal ulceration overlying a papillary dermal infiltrate of
atypical, large, dyshesive cells. The cells had abundant eosinophilic cytoplasm and convoluted nuclei
with red nucleoli. Some cells were multinucleated. Immunohistochemical stains showed that the
histiocytes were positive for CD1a and S-100 protein.
In summary, the morphologic characteristics of the skin lesions in this boy were consistent with
Langerhans’-cell histiocytosis.An extensive evaluation failed to disclose evidence of infection, and
since he remained clinically well, we felt that single-organ–system Langerhans'-cell histiocytosis was
the most likely diagnosis in this case.
The patient was treated conservatively and a surveillance follow up was scheduled.
Notes:
P7.
Integrated study of dermal CD11b+ mononuclear phagocytes including conventional
and monocyte-derived dendritic cells and macrophages present in the mouse skin
Samira Tamoutounour, Martin Guilliams, Frederic Montanana Sanchis, Dora Terhorst, Camille
Malosse, Marc Dalod, Bernard Malissen and Sandrine Henri
Centre d'Immunologie de Marseille-Luminy, Université de la Méditerranée, INSERM U1104, CNRS
UMR6102, 13288, Marseille, France
The mononuclear phagocyte system functions in the innate immune response, in support of the
adaptive immune response and in the maintenance of tissue homeostasis. In the dermis, the lack of
markers permitting the unambiguous identification of macrophages and of conventional and monocytederived dendritic cells complicates the understanding of their contribution to skin integrity and to
immune responses. Recent studies have identified several DC subsets within the mouse skin.
CD11b+MHCII+ cells were found to constitute the predominant myeloid cell type present in the steady
state dermis. Using a panel of markers and appropriate knock-out mouse models, we successfully
identified each cell types within CD11b+MHCII+ cells and studied their origin and their transcriptomic
signatures. We also assessed their migratory and T-cell stimulatory properties. We followed the
monocyte differentiation once they extravasated into the dermis. Moreover, we analyzed the impact of
microbiota on their development and their contribution to skin inflammation during contact
hypersensitivity. Therefore our work provides the first fine-grained description of the DC and
macrophage subsets found in the mouse skin and showed that functional specialization exists among
them.
Notes:
P8.
Could dermal dendritic cells play a role in the development of neutralizing anti-drug
antibodies?
Christina Hermanrud, Anna Fogdell-Hahn
Karolinska Institutet, Stockholm
Interferon beta (IFN-β) is used as a first-line treatment of multiple sclerosis (MS) and can provoke
unwanted immune responses leading to the development of neutralizing anti-drug antibodies (NAbs).
The presence of NAbs is associated with loss of effectiveness of the drug. NAbs develop in 6-47% of
the IFN-β treated MS patients depending on what preparation is used. The mechanisms regulating the
triggering of immunity to antigens administered through the skin are still relatively unexplored and
have previously not been investigated in the context of NAb development, but there are indications
that the route of administration is an essential risk factor. The objective of this project was to
investigate the prevalence of NAbs against IFN-β preparations in 1071 Nordic MS patients analyzed
by our routine NAb testing laboratory during 2011 to 2012, and determine the effects of injection site
(subcutaneous (s.c) versus intramuscular (i.m) injection) on risk for NAbs. The overall prevalence of
NAbs to IFN-β was 19%. IFN-β administered once a week i.m. gave rise to NAb production in 6% of
the patients whereas s.c administration every third day or every other day resulted in NAb production
in 21% and 43% of the patients, respectively. As previous studies have shown; s.c administration of
IFN-β seems more immunogenic than i.m. and higher administration frequency seems to correlate
with increased incidence of NAb development. This supports the indication that the immunology of the
skin has considerable impact on NAb development and thus a potential role of skin dendritic cells.
Notes:
P9.
Casein kinase II regulates the recycling of the DC/LC antigen receptor DEC205 from
MHC class II+ compartments back to the cell surface
Rainer Koch, Sabine Ring, Alexander H.Enk, Karsten Mahnke
Ruprecht-Karls-University Heidelberg, Dept. of Dermatology, Im Neuenheimer Feld 440,D 69120
Heidelberg
The intracellular trafficking of the dendritic cell (DC) antigen receptor DEC205 is guided by an
intracellular domain, that routes antigens into MHC-II+ compartments. This domain contains a putative
casein kinase II (CKII) phosphorylation site and we asked whether this site is involved in intracellular
targeting of DEC205 to late endosomes (LE). We generated fusionreceptors containing the HuIgGbinding, extracellular domain of human CD16 and the intracellular DEC205 domain (CD16:DEC) and
established stably transfected the antigen presenting cell line DCEK. In pulse – chase experiments we
incubated the celllines with HuIgG on ice for 1h, followed by a chase at 37°C for 20 to 60 min. We
show, that CD16:DEC transfected cells bind and endocytose HuIgG efficiently, and after 30 min many
vesicles start to fuse with Rab7+ LE. These vesicles were also positive for CKII indicating a role of
CKII in DEC205 trafficking. When we applied the CKII inhibitor TBB, endocytosis of HuIgG by
CD16:DEC was not affected, but instead the recycling of the DEC receptor from LE to the cell surface
was blocked. Moreover, inhibition of CKII also lead to a prolonged half-life of endocytosed HuIgG
within LE and in antigen presentation assays with HuIgG-specifc T-cells, TBB-treated DCEK cells
induced reduced T cell proliferation as compared to controls. Finally we confirmed our results by
deleting the CKII site in chimeric CD16:DEC receptors by site-directed mutagenesis yielding
CD16:DEC CKII receptors. These receptors accumulated in Rab7+ LE and showed severely reduced
expression on the cell surface. Thus this data indicate that phosphorylation of the intracellular domain
of the DEC205 antigenreceptor by the CKII is crucial for the transport of its ligands beyond the LE to
MHC class II+ compartments and for the recycling of the DEC205 receptor back to the cell surface.
Further investigations of this pathway may explain the efficiency of antigen targeting to DC in vivo.
Notes:
P10.
Characterization of anti-tumoral immune responses in a mouse model of spontaneous
melanoma
David G Mairhofer1, Vincent Flacher2, Suzie Chen3, Juergen C Becker4 and Patrizia Stoitzner1
1
Department of Dermatology and Venereology, Innsbruck Medical University, Innsbruck, Austria
CNRS UPR9021 Immunologie et Chimie Thérapeutiques, Institut de Biologie Moléculaire et
Cellulaire, Strasbourg, France
3
Chemical Biology, Lab for Cancer Research, Rutgers University, Piscataway, NJ, USA
4
Department of Dermatology and Venereology, Medical University Graz, Graz
2
The transgenic mouse strain tg(Grm1)EPv develops spontaneous melanoma with 6 months of age
due to an over-expression of the metabotropic glutamate receptor (Grm1). In our study we aimed to
analyse immune cells within the tumor mass and in draining lymph nodes to learn more about tumor
immunity.
Flowcytometric analyses of tumor cell suspensions and immunofluorescence stainings of tumor
sections were performed. T cell responses were investigated in vitro and in vivo with proliferation and
IFN-gamma secretion assays.
We detected an infiltration of T cells into the tumors when compared to healthy skin. Most infiltrating T
cells were CD4+ rather than CD8+, and included regulatory T cells. In regard to dendritic cells (DC),
we found infiltrating plasmacytoid DC in tumor cell suspensions. Langerhans cells (LC) were depleted
from skin with melanoma, while the Langerin- dermal DC population was enlarged. All tumors
contained a prominent population of myeloid-derived suppressor cells (MDSC). Within lymph nodes
draining tumors, CD4+T cells were decreased in percentages and some expressed FoxP3, thus
representing regulatory T cells. The population of MDSC was increased indicating an
immunosuppressive milieu in the lymph nodes. In addition, we detected less Langerin+ dermal skin
DC and LC in lymph nodes of tumor-bearing mice. Some CD8+T cells were activated and produced
IFN-gamma upon restimulation in vitro. When we adoptively transferred transgenic CD8+ T cells
specific for the melanoma antigen gp100, they proliferated in lymph nodes and proliferation could be
enhanced by intradermal injection of TLR-3 ligand poly I:C plus an antibody against CD40.
In summary we observed that despite some evidence for T cell responses in tumor-bearing mice,
tumors grow progressively in tg(Grm1)EPv mice, most probably due to the lack of and efficient antitumor immune response and the presence of immunosuppressive cells. The exact nature of the tumor
antigen presenting DC subset still needs to be clarified.
Notes:
P11.
EFdA, a reverse transcriptase inhibitor, potently blocks HIV-1 ex vivo infection of
Langerhans cells within epithelium
Takamitsu Matsuzawa1, Tatsuyoshi Kawamura1, Youichi Ogawa1, Kohji Moriishi2, Yoshio Koyanagi3,
Hiroyuki Gatanaga4, Shinji Shimada1, and Hiroaki Mitsuya5,6
1
Department of Dermatology, University of Yamanashi, Japan
Department of Microbiology, University of Yamanashi, Japan
3
Laboratory of Viral Pathogenesis, Institute for Virus Research, Kyoto University, Japan
4
AIDS Clinical Center, National Center for Global Health and Medicine, Tokyo, Japan
5
Department of Infectious Diseases and Department of Hematology, Kumamoto University School of
Medicine, Japan
6
Experimental Retrovirology Section, HIV and AIDS Malignancy Branch, National Cancer Institute,
National Institutes of Health, Bethesda, Maryland
2
Because of the absence of a prophylactic vaccine, new strategies that reduce sexual transmission of
HIV are urgently needed. A recent clinical trial has shown that vaginal application of tenofovir (TDF); a
nucleotide reverse transcriptase inhibitor (NRTI), reduces the probability of sexual transmission of
HIV-1 by an estimated 39% overall. To deliver and demonstrate more potent anti-HIV activity of
microbicides, we have recently developed a novel NRTI; 4’-Ethynyl-2-Fluoro-2’-Deoxyadenosine
(EFdA). Because Langerhans cells (LCs) are suspected to be initial cellular targets for HIV following
sexual exposure to virus, we compared anti-HIV activity of TDF and EFdA in LCs. Strikingly,
preincubation with EFdA (100-5000 nM) completely blocked HIVBa-L infection in monocyte-derived
LCs (mLCs) without cytotoxicity, while preincubation with TDF partially inhibited HIVBa-L infection in
mLCs, and the mean inhibition values with 100, 1000, or 5000 nM TDF was 28%, 37%, and 59%,
respectively. In addition, in an ex vivo skin explant model, when skin explants were preincubated with
each drug, similar blocking effects were observed for epidermal LCs. Moreover, preincubation of
mLCs as well as skin explants with EFdA (100 - 5000nM) completely blocked subsequent HIV
transmission from LCs to cocultured CD4+ T cells, while TDF partially inhibited the transmission.
Similar blocking effect of EFdA was observed when LCs were exposed to R5 HIV primary isolates; JRFL and AD8. In addition, although complete blocking effect of EFdA against HIVBa-L infection in mLCs
persisted for 3 days, TDF gradually lost the anti-HIV activity depending on culture periods. Taken
together, our data suggest that EFdA may be useful as a topical microbicide to block sexual
transmission of HIV.
Notes:
P12.
Differential cytokine requirements for oxazolone and DNCB for allergen-induced
Langerhans’ cell migration
Aleksandra Metryka1, Laura H Eaton1, Ian Kimber1, Ruth A Roberts2 and Rebecca J Dearman1
1
2
Faculty of Life Sciences, The University of Manchester, Manchester, M13 9PL, UK
AstraZeneca, Safety Assessment Alderley Park, Macclesfield, Cheshire, SK10 4TG, UK
We have shown previously that Langerhans’ cell (LC) migration following exposure to oxazolone, a
potent contact allergen, requires two independent cytokine signals from tumour necrosis factor (TNF)α and interleukin (IL)-1β. Moreover, other stimulators of LC migration, including skin irritants are
similarly dependent on TNF-α. However, we have demonstrated recently that 2,4-dinitrochlorobenzene
(DNCB) is able to induce LC migration in the absence of TNF-α signalling. Here we examine further
the mechanisms through which DNCB mobilises LC.
Mice deficient in TNF-α receptor 2 (TNFR2-/-; the sole TNF-α receptor expressed by LC), or wild-type
controls, were exposed topically to the contact allergens: paraphenylenediamine, 1-fluoro-2,4dinitrobenzene (DNFB), DNCB, or to the skin irritant sodium lauryl sulfate. The density of LC was
determined by staining for MHC class II. All chemicals caused significant LC migration in wild-type
animals (between 20 and 30% reduction), however, only members of dinitrohalobenzene family
(DNCB and DNFB) were able to trigger LC mobilisation in TNFR2-/- mice (20-40%).
Subsequently, the kinetics of expression of cutaneous cytokines following treatment of wild-type mice
with either oxazolone or DNCB was examined by RT-PCR. Exposure to oxazolone and DNCB
upregulated the expression of TNF-α, IL-1α and IL-17F and down-regulated mRNA for IL-18 and IL33. However, higher levels of IL-1β (~50 fold versus 5 fold) and CXCL-1 (~120 fold versus 20 fold)
were induced by DNCB compared with oxazolone.
These data reveal that, unlike other inducers of LC migration, dinitrohalobenzenes are capable of
stimulating epidermal LC migration independently of TNF-α. The PCR results demonstrate, that at
concentrations previously inducing similar proliferation levels in the draining lymph node, DNCB and
oxazolone differentially activate IL-1β and CXCL-1 expression. Considering that CXCL-1 expression is
likely to be secondary to IL-1β production, we propose that the TNF-α-independent LC migration
induced by dinotrohalobenzenes is primarily driven by the more vigorous expression of IL-1β.
Notes:
P13.
Revisiting the origin of Langerhans cell histiocytosis: mutated BRAF gene identifies
potential circulating precursor cells
Paul Milne, Venetia Bigley, Naomi McGovern, Matthew Collin
Human Dendritic Cell Lab, Newcastle University Medical School, Newcastle upon Tyne, UK
Langerhans cell histiocytosis (LCH) is a rare, potentially fatal disease caused by clonal proliferation of
langerin+ CD1a+ dendritic cells (DCs), which form lesions in various organs. The cell of origin of LCH
is unknown, though expression of langerin implicates the Langerhans cell (LC). This link is questioned
by numerous recent observations: i) LCH cells have a gene expression profile distinct from LCs ii)
LCH can occur in organs usually devoid of LCs iii) Presence of a normal dermal DC which expresses
langerin+CD1a+, but is phenotypically, functionally and homeostatically distinct from LCs. It is
therefore possible that, similarly to normal tissue DCs, LCH cells derive from circulating precursors
rather than tissue-resident LCs.
60% of LCH biopsies contain an activating V600E mutation in the BRAF gene. Detection of mutated
BRAF in peripheral blood mononuclear cells (PBMC) and bone marrow (BM) is a potential means of
identifying precursors of LCH.
PBMC from two V600E+ multi-system LCH patients were FACS sorted to obtain pure populations of
monocyte, DC and lymphocyte subsets. BM CD34+ progenitors were sorted into five subsets. An
optimised TaqMan Gene Expression Assay was used to perform qPCR for BRAFV600E mutation on
gDNA.
In both patients, V600E mutation was detected in CD14+ ‘classical’, CD16+ ‘non-classical’ monocytes
and CD11c+ myeloid DCs. Peripheral blood T lymphocytes and plasmacytoid DCs were negative. In
bone marrow, V600E mutation was seen in myeloid but not lymphoid progenitors. The presence of
BRAF V600E in peripheral blood monocytes and myeloid DCs is a novel finding, implicating these
cells in the pathogenesis of LCH lesions. The finding of the mutation among BM progenitor cells
shows for the first time that multi-system LCH is a haematopoietic disorder.
Through prospective studies we aim to test the prognostic significance of these findings and validate
the test for diagnostic and disease monitoring purposes.
Notes:
P14.
The role of langerin-positive skin dendritic cells in skin carcinogenesis
Daniela Ortner1, Christoph Tripp1, Nicole Amberg2, Maria Sibilia2, Björn E. Clausen3 and Patrizia
Stoitzner1
1
Department of Dermatology and Venereology, Innsbruck Medical University, Innsbruck, Austria
Department of Medicine I, Institute for Cancer Research, Medical University of Vienna, Vienna,
Austria
3
Institute for Molecular Medicine, University Medical Center of the Johannes Gutenberg-University,
Mainz, Germany
2
Due to their localization skin dendritic cells (DC) are the first antigen presenting cells to get in contact
with transformed keratinocytes forming squamous cell carcinoma (SCC) and basal cell carcinoma
(BCC). It is therefore critically important to understand how skin DC influence immune effector cells in
skin prone to carcinogenesis and in fully developed tumors.
We used two different mouse skin cancer models: (i) a cutaneous two-stage chemical carcinogenesis
model forming SCC which we performed on LangerinDTR:EGFP mice; (ii) a SmoothenedM2
transgene mouse model (Rosa26SmoM2YFP) crossed to K5-CreERT and LangerinDTR:EGFP
(SmoM2LangerinDTR) mice to induce BCC in the absence of langerin-positive DC.
So far experiments showed that depletion of langerin-positive DC prior to the induction of SCC led to a
faster tumor onset and higher number of tumors. Quantitative PCR analysis of treated skin
demonstrated a decrease in IL-12p35, IL-23 and TNF-α expression in the absence of langerin-positive
DC two days after a single application of DMBA. SmoM2LangerinDTR mice depleted of langerinpositive DC exhibited increased tumor load compared to non-depleted mice. Taken together, our first
findings suggest that langerin-positive skin DC are required for immunosurveillance of non-melanoma
skin cancer.
Notes:
P15.
HIV infections in novel CD1a cells from human vaginal mucosa and epidermal
Langerhans cells
Victor Peña-Cruz, Rahm Gummurulu and Manish Sagar
Boston University, Boston Massachusetts, USA
There is great interest in demonstrating HIV infections in human vaginal mucosa to better understand
mechanisms of mucosal HIV acquisition. While skin derived Langerhans cells (LCs) and monocyte
derived dendritic cells (mDCs) have been used as models of the potential earliest target cells present
in mucosal tissue, we examined cells present in vaginal mucosa.
Skin and vaginal cells were obtained from women without cancer undergoing elective reduction
mammoplasties and vaginal repairs respectively. Cells were isolated using discontinuous density
gradients and anti-human CD1a magnetic beads. Cells were phenotyped using flow cytometry and
electron-microscopy (EM). HIV infections were monitored by assessing infectious virus production on
TZM-bl cells.
While both skin epidermis and vaginal epithelium cells expressed CD1a, skin LCs showed a
significantly higher surface expression of Langerin compared to the vaginal cells. In situ EM
examination showed that the presence of Birbeck Granules (BGs) in the skin but, no BGs were evident
in vaginal tissue. BGs were also not observed in pelleted CD1a positive vaginal cells. Diverse HIV-1
strains did not replicate in skin derived LCs, but showed relatively low replication levels in the vaginal
CD1a positive BG negative cells. HIV-2 replicated in cells from both tissues, although replication was
relatively higher in the vaginal as compared to the skin cells.
Our investigations demonstrate a novel vaginal epithelium cell type that expresses CD1a, low levels of
Langerin, and contains no BGs. Compared to skin LCs, the vaginal CD1a positive BG negative cells
support more robust replication with HIV-1 and HIV-2. Our study suggests that these novel vaginal
cells may be some of the earliest infected cells during mucosal HIV transmission.
Notes:
P16.
Distinct molecular signature of human skin Langerhans cells denotes critical
differences in cutaneous dendritic cell immune regulation
Marta E Polak1, Stephen M Thirdborough2, Chuin Y. Ung1, Tim Elliott2, Eugene Healy1, Tom C
Freeman3, Michael R Ardern-Jones1
1
Clinical and Experimental Sciences, Sir Henry Wellcome Laboratories, Faculty of Medicine,
University of Southampton, Southampton, UK.
2
Cancer Sciences, Faculty of Medicine, University of Southampton, Southampton, UK
3
The Roslin Institute and Royal (Dick) School of Veterinary Sciences, University of Edinburgh, United
Kingdom
Langerhans cells (LCs) are dendritic cells (DCs) residing in the epidermis, which demonstrate high
potential to activate T lymphocytes, but due to their relative low frequency, studies of primary human
LC biology are limited. We have obtained complete transcriptome data of human skin migratory LCs
and myeloid dermal DCs (DDCs), before and during the time course of stimulation with TNF-alpha
(pro-inflammatory epidermal cytokine). Genome-wide comparison of the transcriptional profiles of
human skin migratory CD1a+ LCs and CD11c+ dermal dendritic cells (DDCs) demonstrated striking
differences between these ‘dendritic cell’ types. Although LCs conformed to the phenotype of
professional antigen presenting cell (high expression of HLA allele, CD40, CD80, CD86), they differed
distinctively from classical myeloid DCs (including preferential expression of 625 genes (p<0.05) in LC
and 914 genes (p<0.05) in DDC). Analysis of the temporal regulation of molecular networks activated
during stimulation with TNF-α identified 6 clusters of co-dependent genes unique to DDCs and 12
clusters of co-dependent genes unique to LC (p<0.05 - p<0.0001). Whereas gene expression in DDCs
recapitulated the classical DCs function, including surface receptors (CD163, MRC1, Fc-gamma R,
FXIIIa, C-type lectins), soluble immune mediators (IL1a, IL1b, CCL2, CCL17, CCL18, CCL22) and
matrix metaloproteinases (MMP1, MMP3, MMP9), LC preferentially up-regulated genes involved in
cell membrane re-organisation (SYNPO, ACTB) and antigen acquisition and processing (SEPT11,
CAV-1, PSMD14). Functional interference with caveolin abrogated LCs superior ability to crosspresent antigens to CD8+ T lymphocytes, highlighting the importance of these networks to biological
function. The molecular signature of LCs highlights the distinctiveness of this cell population and
underscores their role as tissue gatekeepers, utilising efficiently membrane trafficking as a part of their
role in pathogen surveillance and monitoring of the epidermal microenvironment.
Notes:
P17.
Poly(I:C) induces Th1 cytokines and IFNbeta in monocyte-derived Langerhans-like cells
through TLR3
André Said, Monika Schäfer-Korting, Günther Weindl
Langerhans cells (LC) might contribute to tolerance towards bacterial commensals, but are ideally
positioned in the epidermis to induce anti-viral immunity. Here we investigated the immune responses
of human immature monocyte-derived DC (MoDC) and LC-like cells (MoLC) upon stimulation with
different TLR ligands in the presence or absence of pro-inflammatory cytokines. MoLC were
distinguished from MoDC by the expression of E-Cadherin, Langerin, TROP-2 and Axl. Bacterial TLR
ligands selectively activated MoDC and induced the release of distinct cytokines to prime a Th1 or
Th17 response. In contrast, MoLC weakly upregulated CD83 and CD86 expression in response to
bacterial antigens, whereas activation under inflammatory conditions (IL-1beta, TNF) strongly
enhanced a phenotypically mature state and increased the release of IL12p70, IL-23 and IL-6.
Moreover, poly(I:C), a molecular pattern associated with viral infections, led to upregulation of CD83
and CD86 surface expression and release of IL-12p70 in stimulated MoLC. The addition of proinflammatory cytokines further enhanced the production of IL12p70 and IFNgamma. Additionally,
poly(I:C)-stimulated MoLC showed increased levels of IFNbeta mRNA, whereas IFNalpha and
IFNbeta were upregulated in MoDC. Pretreatment with cholorquine abrogated IFN induction,
suggesting a TLR3-dependent activation of both, MoLC and MoDC by poly(I:C). Taken together, the
obtained results indicate that human Langerhans cells, besides their tolerogenic function to bacterial
commensals in healthy human skin, might contribute to an adaptive immune response towards
bacterial pathogens under inflammatory conditions. Furthermore, we suggest a critical involvement of
MoLC and MoDC in innate defense against viral pathogens, potently in immunologic synergy with
pDC, the main producers of type I IFNs. In addition, the altered cytokine secretion favors MoLC as
potent inducers of a Th1 response against viral pathogens under inflammatory conditions.
Notes:
P18.
Human Langerhans cells play a crucial role in CCR5 tropic selection of HIV-1 variants
during sexual transmission
Ramin Sarrami-Forooshani, Annelies W. Mesman, Carla M.S. Ribeiro, Michiel van der Vlist and
Teunis B. H. Geijtenbeek
Meibergdreef 9, 1105 AZ Amsterdam, The Netherlands
The HIV pandemic is mainly fueled via sexual transmission. In addition to CD4, HIV-1 requires
coreceptors of the CC or CXC chemokine families for productive infection. Based on coreceptor
tropism there are two main HIV-1 variants, X4 and R5 which use CXCR4 or CCR5 for the infection,
respectively. During HIV transmission only R5-HIV-1 variants can be transmitted however the
mechanism of this selection is yet to be explained. During sexual transmission, Langerhans cells (LCs)
are the first component of the immune system which encounter the virus. Under inflammatory
conditions LCs become infected and transmit HIV-1 to T cells. To investigate the role of primary LCs
on the selection of R5-HIV-1 variants, we set up different transmission assays including ex vivo human
skin explant model which more closely resembles the physiological condition and in situ exposure. Our
data showed an efficient transmission of R5 viruses compared to X4 ones. Interestingly LCs showed a
productive infection with X4-HIV-1 variants. Treatment of LCs with HIV-1 inhibitors revealed that de
novo infection of LCs is required for HIV transmission (infection in cis). When LCs were exposed to
HIV-1 after migrating out from the skin (migratory LCs), not only they were infected but also
transmitted the X4-HIV-1 variants. Considering the activation state as the main difference between
immature and migratory LCs, epidermal sheets were treated with different stimuli. Notably the
immature LCs that were stimulated with TNF-alpha could transmit the X4-HIV-1 variants. Our data
reveals the important role of LCs in R5-HIV-1 selection. Furthermore, it suggests that inflammatory
conditions affect this selection by facilitating X4-HIV-1 transmission. Our study demonstrates that the
role of LCs on this selection is not due to lack of infection with X4 but possibility through existence of
inhibitory factors that may block transmission of X4-HIV-1 variants.
Notes:
P19.
Targeting the glutamate pathway: a potential approach for immunotherapy in
melanoma
Sandra C. Schaffenrath1,2, Daniela Ortner1, Suzie Chen3, Nikolaus Romani1,2 and Patrizia Stoitzner1
1
Department of Dermatology and Venereology, Innsbruck Medical University, Austria
Oncotyrol – Center for Personalized Cancer Medicine, Innsbruck, Austria
3
Division of Medical Oncology, University of Medicine and Dentistry of New Jersey, USA
2
Glutamate is a key compound in cellular metabolism as well as the most abundant neurotransmitter in
the CNS and exerts its effect by binding to ionotropic- and metabotropic glutamate receptors. More
than 60% of human melanoma samples are positive for ectopic overexpression of metabotropic
glutamate receptor-1 (Grm1) which is absent in melanocytes. Aberrant expression of Grm1 in
melanocytes induces spontaneous melanoma development and release of elevated levels of
glutamate accelerates tumor growth. In the tg(Grm1)EPv mouse model overexpression of Grm1 in
melanocytes leads to spontaneous tumor formation in hairless skin regions. In this study with
investigated how Riluzole, a pharmacological inhibitor of glutamate release, affects growth of
melanoma cells. Moreover, we want to know if induction of tumor cell death changes immune
responses against melanoma.
In vitro experiments demonstrated that various glutamate pathway inhibitors induced apoptotic cell
death in Grm1 overexpressing cell lines in a dose-dependent way. In addition to our in vitro findings
we observed that Riluzole treated mice showed tumor growth delay compared to vehicle control over a
period of 2-3 month. Flowcytometric analyses revealed that Riluzole had minor effects on the
percentages of infiltrating immune cells in in the tumor as well as tumor draining lymph nodes.
Currently, we are investigating the consequences of Riluzole treatment on phenotype and function of
immune cells like dendritic cells (DC), natural killer cells (NK) and T cells.
Our findings so far suggest that growth of melanoma can be inhibited by interfering with the glutamate
pathway, however, cell death might be non-immunogenic. Our observations implicate glutamate
signaling as new therapeutic target for treatment of melanoma.
Notes:
P20.
Novel hollow microneedle technology for depth controlled microinjection-mediated
dermal vaccination: a study with polio vaccine in rats
Koen van der Maaden1 *, Bas Trietsch2 *, Pim Schipper1, Juha Monkare1, Heleen Kraan3, Eleni Maria
Varypataki1, Stefan Romeijn1, Raphäel Zwier4, Heiko van der Linden2, Gideon Kersten1,3, Thomas
Thomas Hankemeier2, Wim Jiskoot1, Joke Bouwstra1
1
Division of Drug Delivery Technology, Leiden Academic Centre for Drug Research (LACDR), Leiden
University, P.O. Box 9502, 2300 RA Leiden, the Netherlands
2
Division of Analytical Biosciences, Leiden Academic Centre for Drug Research (LACDR), Leiden
University, P.O. Box 9502, 2300 RA Leiden, the Netherlands
3
Institute for Translational Vaccinology (Intravacc), P.O. Box 450, 3720 AL Bilthoven, the Netherlands
4
Fine Mechanical Department, Leiden Institute of Physics (LION), Leiden University, P.O. Box 9502,
2300 RA Leiden, the Netherlands
* Authors contributed equally
Conventional intramuscular or subcutaneous injections of vaccines cause pain and stress, carry the
risk of infection and need trained personnel. By only piercing the top layers of the skin, microneedles
can be used for minimally invasive, pain free vaccine delivery to an immunologically active site.
Here we present a cheap and fast method to produce hollow microneedles from fused silica capillary
with a defined and tunable inner diameter. The microneedles were used in combination with an
electromagnetic applicator to control the insertion speed (1-3 m/s), depth (0-1000 µm), and angle (10°90°) for painless intradermal vaccine delivery.
Hollow microneedles with an inner diameter of 20 µm were used to immunize rats with an inactivated
poliovirus vaccine by an intradermal microinjection of 9 µL at a depth of 300 µm and an insertion
speed of 1 m/s. Intradermal microinjection induced comparable immune responses to conventional
intramuscular injection, demonstrating the potential of microneedles for pain free, minimally invasive
intradermal vaccination.
Notes:
P21.
CD14+ DDCs take the stage in allergen-specific intradermal immunotherapy
Na Luo1, Yuri Souwer1, Fong Lin1, Ghaith Bakdash1,Toni M van Capel1, Martien L Kapsenberg1,
Marcel BM Teunissen2 and Esther C de Jong1
1
Department of Cell Biology & Histology, Academic Medical Center, University of Amsterdam,
2
Department of Dermatology, Academic Medical Center, University of Amsterdam, Amsterdam, The
Netherlands
The most common type of allergy is IgE-mediated type I hypersensitivity which affects approximately
25% of the population in industrialized countries. Allergen specific immunotherapy (SIT) currently is
the only curative treatment of mono allergies, however clinical improvement is only seen after
treatment for five years. The efficacy of SIT is thought to be based on induction of regulatory T cells
that suppress disease-associated Th2 cells. The typical route of administration for SIT is
subcutaneous injection (SCIT). The skin is an interesting site for vaccination as it is equipped with an
extensive network of different dendritic cell (DC) types: epidermal Langerhans cells (LCs) and CD1a+
and CD14+ dermal dendritic cells (DDCs). It is unclear to what extent these individual DC types
contribute to SCIT. To study the possible roles of different DC types in SCIT we used a human skin
explant model that closely resembles physiological conditions. After intradermal delivery of fluorescent
labelled allergen, biopsies were taken and cultured for 1-3 days. Cells that migrated out of the skin
were analyzed. This showed that allergen was preferentially taken up by CD14+ DDCs. Since the
active form of vitamin D3 (VitD) is a potent immunosuppressive drug and its effects are mediated in
part through DCs that promote the development of regulatory T cells, we administered VitD in our
system. Injection of VitD selectively enhanced the migration of CD14+ DDCs. Moreover, these
migrating DDCs induced T cells with suppressive activity, characterized by expression of Foxp3, and
abolished T cell IFN-γ production. We show that allergen injected in the skin is taken up by CD14+
DDCs and injection of VitD in the skin increases the migratory capacity of CD14+ DDCs which induce
regulatory T cells. Our results suggest that co-administration of allergen and VitD during SCIT could
greatly improve the efficacy of this treatment.
Notes:
P22.
Dengue receptor CLEC5A induces immune responses in skin Langerhans cells
Joris K. Sprokholt, Sonja I. Gringhuis, Teunis B.H. Geijtenbeek
Amsterdam, the Netherlands
Dengue is a mosquito-transmitted disease caused by dengue virus, which infects 400 million people
annually. The disease is characterized by massive cytokine induction and vascular permeabilization,
causing internal bleeding. The C-type lectin receptor CLEC5A is a receptor for dengue virus and
thought to be involved in the cytokine ‘storm’. Little is known about the expression and functionality of
CLEC5A on skin Langerhans cells (LCs), which are thought to be the first cells to encounter the virus
after a mosquito bite. Here we show that CLEC5A is highly expressed on immature epidermal LCs.
Notably, activation of CLEC5A on primary LCs using an agonist antibody leads to profound immune
responses such as cytokine secretion, maturation and modulation of TLR signaling. The production of
inflammatory cytokines and chemokines induced by CLEC5A consists of IL-1β, IL-6, IL-23, TNF-α,
CLL3 and CLL4. Interestingly, CLEC5A activation modulates TLR-induced immune responses by
increasing TLR-induced cytokines and by inducing additional cytokines such as IL-12. In addition,
CLEC5A activation induces maturation of primary LCs by upregulating CD80 and CD86. We are
currently confirming these results using dengue virus. Taken together, LCs may be the first cells to
start the cytokine ‘storm’ and inhibiting the function of LCs might be a key strategy to combat the
disease at an early stage of infection.
Notes:
P23.
Polycomb Group Protein Ezh2 Regulates Integrin-Dependent Leukocyte Migration and
Methylation of Talin
M. Gunawan, N. Venkatesan, W.H. Neo, J. Li, J.T. Loh, J.F. Wong, T. Guo, C.E. See, S. Yamazaki,
K.C. Chin, L.G. Ng, K.Z. Sze, F. Ginhoux, I-H. Su
Division of Molecular Genetics and Cell Biology, Division of Structure Biology and Biochemistry,
School of Biological Sciences, College of Science, Nanyang Technological University
We previously identified a novel cytosolic role for the histone methyltransferase Ezh2 in the regulation
of cell signaling and in the control of receptor-stimulated actin polymerization. This unexpected
cytoplasmic role for Ezh2 is likely to profoundly impact on key leukocyte functions by modulating signal
transduction pathways in these cells, but the molecular mechanism that underpins the extra-nuclear
functions of Ezh2 has until now remained unclear. In the current report, we present the first
experimental evidence that talin is a cytosolic substrate of Ezh2, and we provide novel mechanistic
insights into the influence of Ezh2 on leukocyte adhesion dynamics and migratory behavior both in
vitro and in vivo.
Here we show that Ezh2 critically regulates integrin signaling and governs the adhesion dynamics of
neutrophils and dendritic cells (DCs). Ezh2 deficiency impaired integrin-dependent transendothelial
migration of innate leukocytes and restricted disease progression in an experimental autoimmune
encephalomyelitis model of multiple sclerosis. Ezh2 was necessary for methylation of the talin protein
that links integrin molecules to the actin-cytoskeleton, and Ezh2 interactions with the Vav1 guanine
nucleotide exchange factor were essential for the regulation of adhesion structure turnover in DCs.
The influence of Ezh2 on leukocyte migration also extends to the steady-state trafficking of
Langerhans cells through a continuous physical barrier (basement membrane) to LN. Further
investigation will now be required to assess physiological outcomes of this regulation. Our data
demonstrate for the first time that leukocyte migration and adhesion dynamics are critically regulated
by polycomb group protein Ezh2 and talin methylation.
Notes:
P24.
Unique role of Cbfb2 variant in Langerhans cell development
Mari Tenno, Yoshinori Naoe, Sawako Muroi and Ichiro Taniuchi
Laboratory for Transcriptional Regulation, RIKEN, Research Center for Allergy and Immunology,
Yokohama, Kanagawa, Japan
Langerhans cells have been known to be a unique and important subset of dendritic cells in the
epidermal layer of the skin; its developmental pathway is however not fully understood. In mammals,
Cbfb protein, known as an essential subunit of Runx transcription factor complexes, are present as
two RNA splicing variants, Cbfb1 and Cbfb2. These two variants differ in their C-terminal amino acid
sequences, suggesting a possibility that each isoform may have distinct physiological function. To
address in vivo roles of individual variant, we have generated mutant mouse strain lacking either
Cbfb1 or Cbfb2. Here we show that a loss of Cbfb2 variant results in a defect in Langerhans cell
development. In the ear epidermis of adult Cbfb2-deficient mice, no mature Langerhans cells
expressing Langerin and EpCAM were detected. Instead, there were a few larger cells with
characteristics of immature Langerhans precursors, such as the dendritic morphology and the
expression of MHC class II and CX3CR1. Analysis of bone marrow chimera showed that these
Langerin+EpCAM+classII+ cells were radio-resistant residential cells that could not be replaced by
donor cells. Analyses of ontogeny of Langerhans cell development around neonatal stage revealed
that seeding of CD45+ early progenitor cell to the skin occurred normally in Cbfb2-deficient mice.
However, the next transition into Langerin+EpCAM+ stage, which is known to require for TGFb signals
around day 3 after birth, was severely impaired in the absence of Cbfb2. Interestingly, although
immature Langerhans precursors disappear in adult epidermis by lack of TGFbR1 signals, our results
indicate that those cells are stably maintained in the Cbfb2-deficient mice, suggesting an unique role
of Runx3/Cbfb2 complexes in regulating Langerhans cell development.
Notes:
P25.
Characterization of dendritic cell subtypes in human melanoma and non-melanoma
skin cancer
Christoph H. Tripp1, Van Anh Nguyen1, Dietmar Heiser1, Barbara Del Frari2, Nikolaus Romani1,3,
Patrizia Stoitzner1
1
Department of Dermatology and Venereology, Innsbruck Medical University, Austria
Department of Plastic, Reconstructive and Aesthetic Surgery, Innsbruck Medical University, Austria
3
ONCOTYROL - Center for Personalized Cancer Medicine, Innsbruck, Austria
2
In the skin, dendritic cells (DC) are the prime cells to induce immune responses against melanoma
and non-melanoma skin cancer (NMSC), such as squamous cell carcinoma (SCC) and basal cell
carcinoma (BCC). Skin tumors are exceedingly common due to cumulative UV exposure and recurrent
in immunsuppressed patients. Little is known about the occurrence, phenotypes and functional
capabilities of tumor-infiltrating DC and their specific role in skin cancer.
The aim of our study is to identify tumor-infiltrating DC subsets, their respective activation states and
expression levels of C-type lectins by extensive FACS analyses. The tumor milieu will be investigated
by analyses of tumor-infiltrating immune cells, such as immunsuppressive tumor-associated
macrophages (TAM) and myeloid-derived suppressor cells (MDSC) as well as effector T and NK cells.
By comparison of cutaneous tumors of melanoma, SCC and BCC we hope to learn more about the
role of human skin DC in cancer. Preliminary data reveal the presence of different DC subtypes and
activation levels in cutaneous metastases of melanoma, SCC and BCC. A substantial proportion of
these DC express DEC-205 which would allow in situ targeting of tumor-infitrating DC. The TLR3
ligand poly(I:C) improves DEC-205 targeting in migratory skin DC, thus rendering it a promising
adjuvant for immunotherapy. Moreover, infiltrates of T cells, NK cells and macrophages were obvious
in all tumor entities. We are currently investigating interactions between these cells by
immunofluorescence stainings of tumor sections.
This study will help to elucidate which DC subtype would be the optimal target for the clinical use of an
“antibody-targeting” immunotherapy.
Notes:
P26.
Activated vaginal Langerhans cells efficiently transmit HIV-1 to T cells
Nienke H. van Teijlingen, Carla M.S. Ribeiro, Elisabeth van Leeuwen, Joris A.M. van der Post, Teunis
B.H. Geijtenbeek
Experimental Immunology & Obstetrics and Gynaecology department, Academic Medical Center,
University of Amsterdam, Amsterdam, The Netherlands
HIV-1 infection is still a global health problem and sexual transmission is the major route of infection.
Development of effective microbicides is urgently needed but little is known about the cellular and
molecular processes in HIV-1 transmission. In sexual transmission Langerhans cells (LCs) are the first
immune cells to encounter HIV-1 and these cells play a central role in the ensuing immune response
but can also transmit HIV-1 to T cells. However, vaginal LC research has been hampered due to
scarcity of tissue and complexity of isolation procedures.
We have successfully developed methods to isolate both immature and migratory LCs from vaginal
mucosa. We have analyzed the expression and function of pattern recognition receptors (i.e. C-type
lectin receptors and toll-like receptors) on these cells and studied the interaction between vaginal LCs
and HIV-1. Migratory vaginal LCs, which highly express LC activation markers, become infected with
HIV-1 and efficiently transmit HIV-1 to T cells.
Thus, vaginal LCs play a remarkable role in HIV-1 infection and further studies on vaginal LCs will
provide valuable knowledge to facilitate the design of preventative or therapeutic strategies to combat
HIV-1 infection and AIDS. Future studies will focus on the influence of vaginal LC activation by
bacterial vaginosis, an imbalance of vaginal flora, which is associated with increased HIV-1
susceptibility.
Notes:
P27.
Innate dendritic cell sensing of opsonized HIV-1 to activate Th17 cells
Wilfried Posch1, Andrea Schroll2, Hubert Hackl3, Cornelia Lass-Flörl1, Gianfranco Pancino4, Asier
Saez-Cirion4, Zlatko Trajanoski3, Teunis Geijtenbeek5, Günter Weiss2, Doris Wilflingseder1
1
Division of Hygiene and Medical Microbiology, Innsbruck Medical University, Schöpfstrasse 41/R311,
Innsbruck, Austria
2
Department of Internal Medicine VI, Clinical Immunology and Infectious Diseases, Innsbruck Medical
University, Anichstr. 35, Innsbruck, Austria
3
Division of Bioinformatics, Biocenter, Innsbruck Medical University, Innrain 80-82, Innsbruck, Austria
4
Unité de Régulation des Infections Rétrovirales, Institut Pasteur, 25 Rue du Docteur Roux, 75724
Paris, France
5
Center for Experimental and Molecular Medicine, AMC, University of Amsterdam, Amsterdam,
Netherlands
Early on in HIV-1 infection, gut Th17 cells are massively depleted leading eventually to compromised
intestinal barrier function and excessive immune activation. In contrast, the functional Th17 cell
compartment of the gut is well-maintained in non-pathogenic SIV infection as well as HIV-1 long-term
non-progressors. Here, we show that dendritic cells (DCs) exposed to HIV-1 bearing high surface
complement levels, following incubation in plasma from HIV-infected individuals, secreted significantly
higher concentrations of Th17-polarizing cytokines compared to DCs exposed to non-opsonized HIV-1
or HIV-1 exhibiting low levels of surface complement following opsonization in patient plasma. Our in
vitro and ex vivo data, therefore, indicate that complement-opsonized HIV-1 exerts beneficial effects
during HIV-1 infection by simultaneously triggering Th17 expansion as well as stronger CTLs via DCs
as illustrated earlier by our group. Thus, specifically modifying the complement signaling pathway
could strenghten the cellular arm of the immune system and serve as a therapeutic target for HIV-1
infection.
Notes:
P28.
Protease inhibitor from tick saliva acts on human lysosomal enzymes in differentiated
MUTZ-3 cells
Tina Zavašnik-Bergant1, Lenka Grunclová2, Jiří Salát2, Robert Vidmar1, Petr Kopáček2, Boris Turk1
1
Department of Biochemistry, Molecular and Structural Biology, Jožef Stefan Institute, Ljubljana,
Slovenia
2
Institute of Parasitology, Biology Centre of the Academy of Sciences of the Czech Republic, České
Budĕjovice, Czech Republic
By transmitting pathogens ticks represent a severe threat to humans and animals. Furthermore, saliva
of blood-feeding parasites, such as ticks, is a rich source of secretory proteins (sialome). Among them
many have been proposed to help the parasite to overcome the defense response in host skin. Here
the interactions between salivary cystatin-type inhibitor (OmC2 from the saliva of soft tick Ornithodoros
moubata) and host lysosomal enzymes (human cysteine proteases) are described. Antigen-presenting
cells (APC) have the capacity to process foreign proteins into antigenic peptides and trigger specific
immune response. Namely, with the tick inhibitor entering into the endocytic pathway of APC in skin,
the proteolytic ability of host lysosomal proteases (among them cysteine cathepsins S and L,
essentially involved in antigen processing and consecutive MHC II presentation) would be impaired. A
recombinant tick cystatin OmC2 was studied together with differentiated MUTZ-3 cells applied as an in
vitro cell model for dermal APC. First, internalization of the fluorescently labeled tick cystatin was
followed into acidic organelles, where target proteases resided. OmC2 inhibitory effect towards
cysteine cathepsins was tested: (1) with recombinant human cathepsins S and L and (2) in lysosomeenriched lysates from differentiated MUTZ-3 cells after incubation and internalization of tick inhibitor,
respectively. In both cases a decreased specific proteolytic activity of cysteine proteases was
established. Formation of complexes between human lysosomal proteases and tick inhibitor OmC2
was further confirmed with mass spectrometry. Finally, a decrease in IL-12 secretion was observed
when tick inhibitor was internalized into acidic organelles of differentiated MUTZ-3 cells, compared to
non-treated cells. We conclude that tick inhibitor OmC2 could impair the necessary proteolytic events
associated with MHC II antigen presentation in skin APC.
Notes:
97 UPCOMING MEETING
DC2014
The 13th International Symposium on dendritic cells
Tours, France
September 14 – 18, 2014
Organized by
Sebastian Amigorena, Matthew Albert, Jacques Banchereau,
Philippe Pierre & Laurence Zitvogel
www. DC-2014.com
LC2015
14th International Workshop on Langerhans Cells
Kyoto, Japan
November 5 – 8, 2015
Organized by
Kenji Kabashima, Tatsuyoshi Kawamura, Kayo Inaba & Shinji Shimada
www.LC2015.jp
98 The new image processing platform
for skin analysis by TissueGnostics
Superior colour separation
& cell nucleus detection
Epidermis detection
Langerhans Cell detection
Dendrite reconstruction
Distance measurements
of cells within
of cells outside of
the epidermis
www.tissuegnostics.com

- 13th International Workshop on Langerhans Cells

Transcription

Similar documents

Congenital Letterer

Spring `14 Lecture 1 slides

Feel Your Best with the BOOM!

when animal research hits the headlines

Histiocytes and Histiocytosis

[FLY] Immune Support - Deseret Biologicals

Hair Test 74

7th Annual - Feinberg Labs

Ambrotose®—The Missing Link in Human

Dendritic excitability modulates dendritic information

Tumor Antigens and Cancer Vaccines

section home - Centre for Trophoblast Research

Disseminated Human Immunodeficiency Virus 1 (HIV

Y-chromosome descent clusters and male differential reproductive